In the lymph nodes, expressions of both CSCs, and immune suppression markers were upregulated significantly compared with that in primary tumors. The mRNA expression of selected CSCs and immune suppression markers exhibited the highest expression in the level II of metastasis, then declined in the level III and remained constant at a reduced value in levels IV and V of metastases. These results reveal a comprehensive understanding of the unique genetic characteristics associated with metastatic loci and potential routes of lymphatic dissemination of HNSCC, which helps to explain why the level II has a high incidence of lymph node metastasis, and why skip metastasis straight to the level IV or level V is rarely found in the clinic.The progress of Head and Neck Squamous Cell Carcinoma (HNSCC) is dependent on both cancer stem cells (CSCs) and immune suppression. This study was designed to evaluate the distribution of CSCs and the characteristic immune suppression status in HNSCC primary tumors and lymph nodes. A total of 303 lymph nodes from 25 patients, as well as tumor and adjacent normal tissue samples, were evaluated by a quantitative PCR assay of the markers of CSCs and the characteristic immune suppression. Expressions of selected genes in The Cancer Genome Atlas (TCGA) datasets were also analyzed. In the primary tumors, we found that expressions of CSCs markers  Head and Neck squamous cell carcinoma (HNSCC) is ranked as the sixth most common cancer in the world, with almost 600,000 new cases occurring every year  was 57, ranging from 52 to 74. The samples were collected during the surgery and immediately frozen, including cancer tissues, adjacent normal tissues and lymph nodes from the same patient. Adjacent normal tissues were required to be more than 2 cm from the tumor margins in the same patient. The categorization of neck dissection samples was in accordance with the topographic classification of cervical lymph node levels suggested by Gregoire et al.  as IA, IThe patients in one of the following situations were excluded in this study . The final number of HNSCC patients included was 509. The statistical programing software R (version 2.14.1) was used to analyze the datasets with the statistical significance at P < 0.05. The normalized counts  were used to compare the gene expression (RNA-Seq version 2).The selected gene expression was downloaded from mRNA expression detection platform RNA-seq version 2 (level 3) in TCGA data portal  was used to extract the total RNAs of all the acquired samples, and iScriptTMcDNA synthesis kit  was used to reverse transcription (RT). The FastSYBR Green master mix with Rox  was used to perform the quantitative PCR. Fifteen genes were evaluated by quantitative RT-PCR. The primers for (−ΔΔCt). We considered the data was significant at P < 0.05.Data from more than three independent experiments are represented as the mean ± standard deviation (SD). Pared t test was used to do statistical analyses comparing the genes expression between primary and adjacent normal tissues of HNSCC patients, Mann Whitney test was performed to analyze the genes expression in metastatic nodes and non-metastatic nodes and data from TCGA, Kruskal–Wallis test was used to access the expression of metastatic nodes in different levels. Mean-normalized mRNA expression value in non-metastasis lymph nodes were chosen to be control for the entire cohort of lymph nodes. The relative value of each gene was calculated as follow: The ΔΔCt = tumor ΔCt-control ΔCt, fold change of mRNA was obtained as 2n = 25) including age, gender, tobacco and alcohol history, tumor site, and AJCC stage [which was according to AJCC 8th Edition  in the primary tumors of HNSCC. The Clinicopathologic characteristics of TCGA dataset was shown in ALDH1L1, PECAM1, SMAD3, TMEM173, PROM1, and STAT3 were expressed lower, and the rest of the genes were expressed higher in primary tumors than in adjacent normal tissues. Among them, the expression levels of IRF1, IFNAR2, FOXP3, CD47, ALDH1L1, PROM1, EGFR, SOX2, TGFB1, and STAT3 were significant (P < 0.05) between tumors and normal tissues , while the TCGA dataset revealed no significant difference, the expression levels (up or down) of the rest 14 selected genes were consistent with TCGA profiles as validated by real time RT-PCR assay for all recruited patients and a clustering analysis was performed in the primary of HNSCC . It suggested that sex factor might play a role in the formation of CSCs and immune suppression and might be more susceptible to metastasis.To further investigate the above-mentioned gene expression in our HNSCC samples, we validated the mRNA expression using real time RT-PCR detection. The differential levels of mRNA expression (–ΔΔCt) of the 15 genes in each of the 25 individuals HNSCC sample are shown in of HNSCC . Furtherof HNSCC . We alsoCD47, EGFR, FOXP3, IFNAR2, IRF1, SOX2, BMI-1, PROM1, and TGFB1 were upregulated significantly in metastatic nodes when compared to non-metastatic nodes (STAT3 and PECAM1 were decreased in primary but upregulated in lymph nodes (P > 0.05)  , 3. One ic nodes . Finallyic nodes . Also, iic nodes .−) always occurred in an older patient population and their clinical outcomes were unquestionably worse than HPV+ HNSCC, and few HNSCCs are associated with HPV infection. Therefore, our results provide a model to determine gene expression patterns in both the primary tumors and metastatic lymph nodes of HPV− HNSCC.The nodal dissemination of carcinomatous cell is a vital determinant of prognosis in patients with HNSCC, as the overall survival rate of patients with metastatic lymph nodes decreased by a half compared with that without nodal implicated , and TWISTNB (TWIST NEIGHBOR) upregulated in lymph nodes. SOX2, BMI-1, and PROM1 are cancer stem cells-related genes  by EMT (Schlereth et al., It has been reported that ALDH1 has higher activity in the stem cell subclone of leukemia and some solid tumors (Li et al., CD47 is likely involved in the process of evading immunological eradication (Chattopadhyay et al., STAT3 overexpression in metastatic sites may restrain the immune responses to render an immunosuppressive environment (Punt et al., CD47 and FOXP3 were significantly upregulated both in primary site and lymphatic loci, but STAT3, and PECAM1 were only upregulated in lymph nodes. It indicates that there may form a more immunosuppressive environment in the lymph nodes than primary site and may exist an aggressiveness niche to foster metastasis in HNSCC patients.A previous study revealed that male patients may have higher tendency of cancer-associated gene expression in the primary than matched adjacent normal tissue (Shores et al., It have been reported that a relatively constant and sequential route might exist in the lymphatic drainage of the head and neck region Ji, . Level ISMAD3, PECAM1, TMEM173, STAT3, and TWISTNB reveals no statistically significant difference among different groups. Meanwhile, our observation revealed PECAM1 was significantly decreased but it showed no significant difference in TCGA datasets. It may result from that the squamous cell cancer in oropharynx, hypopharynx, and larynx were included in TCGA dataset, and they had an entirely different biological behavior and prognosis from HNSCC. While these findings will require further validation in larger cohorts of patient samples in the future, we believe this first gene expression analysis of cervical lymph nodes in the individual level of cancer patients will provide us an important opportunity to guide the future investigations of HNSCC.The relatively small quantity of lymphatic metastasis samples, the variations of tumor sample purity, and the intratumor heterogeneity limit our current study, especially given that the expression of Through gene expression analyses, we found that the mRNA expression of selected CSCs and immune suppression markers exhibit the highest expression in the level II metastatic lymph nodes, then declined in the level III and remained constant at a reduced value in levels IV and V metastatic lymph nodes. These results help to explain the reason why the level II has a high incidence of lymph node metastasis, and skip metastasis to the level IV or level V is rarely found in the clinic. It will help to increase the understanding of the genetic characteristics associated with metastatic loci and potential routes of lymphatic dissemination of HNSCC, and may aid the clinical diagnosis and treatment of HNSCC.All datasets generated for this study are included in the article/The studies involving human participants were reviewed and approved by Shanghai Ninth People's Hospital Ethical Committee. The patients/participants provided their written informed consent to participate in this study.SQ wrote the paper. ZH, RY and LL carried out experimental studies. LM and SL were involved in statistical analysis. GR and JH modified the paper and designed this study concepts. All authors read and approved the final manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


With an estimated contribution of up to 6% of all acute coronary events, young adults are experiencing an escalating burden and mortality attributable to coronary artery disease (CAD) worldwide. Wellens syndrome, a preinfarction clinical entity with distinctive electrocardiographic (ECG) features and high propensity for extensive anterior wall infarctions, affects about 15% of unstable angina subpopulation. We report challenges and lessons learnt from the first ever documented case of Wellens syndrome in Tanzania. A 30-year-old female of African descent was referred to us from an upcountry zonal referral hospital for etiological determination of chest pain and expert management. Her medical history was unremarkable, and she had no apparent risk factors for CAD. She presented with a 7-day history of ongoing sharp central chest pain that was radiating to the neck and jaws and relieved momentarily by morphine. She had stable vitals with an unremarkable systemic examination; however, a 12-lead ECG revealed deeply inverted T-waves on leads V2 through V4. Based on our extensive history and physical examination we came up with a diagnosis of type B Wellens syndrome with impending anterior wall myocardial infarction. She underwent cardiac catheterization that revealed a nonthrombogenic severe subocclusive (>95%) mid left anterior descending artery stenosis. Subsequently, angioplasty was performed successfully with a resolute integrity stent, and TIMI III flow was achieved. To conclude, despite its relative frequency, physicians’ awareness of Wellens syndrome pathognomonic ECG features is of paramount importance to curb its attributable morbidity and mortality. The dilemma in recognizing this portentous syndrome lies in (1) inadequate physician skills in ECG interpretation, (2) frequent absence of clear ischemia at initial evaluation, (3) frequent manifestation of the signs during pain-free intervals, and (4) the sharing of the characteristic ECG pattern with several other common clinical conditions including hypokalemia, intracerebral hemorrhage, persistent juvenile T-wave inversion, pulmonary embolism, right bundle branch block, or at times it may be just a normal variant.19 With the increasing global prevalence of CAD risk factors in younger populations,10 an increase in presentation of Wellens syndrome at earlier age is foreseeable. We report the experience and lessons learnt from the first ever documented local case of Wellens syndrome in a 30-year-old woman of African origin from northern Tanzania.Wellens syndrome affects all age groups and shares the traditional risk factors with coronary artery disease (CAD), that is, smoking, obesity, dyslipidemia, hypertension, diabetes mellitus, metabolic syndrome, and family history of premature CAD.A 30-year-old female primary school teacher of African descent was referred to Jakaya Kikwete Cardiac Institute from a northern zone referral hospital (540 km away) for etiological determination of chest pain and expert management. She has never smoked cigarettes or used illicit drugs and denied a family history of heart disease. Her medical history was only significant for menorrhagia. She presented to us with a 7-day history of ongoing sharp central chest pain that was radiating to the neck and jaws and relieved temporarily by morphine. The pain woke the patient from sleep and was accompanied by shortness of breath, diaphoresis, and nausea without emesis. She was hospitalized at the upcountry hospital for the initial 3 days of chest pain and was prescribed acetylsalicylic acid 75 mg od (once a day), clopidogrel 75 mg od, atorvastatin 40 mg nocte (every night), and syrup morphine PRN (as needed). Cardiac biomarkers and echocardiography (ECHO) were unremarkable, but her initial ECG (done 7 days prior) showed a strain pattern .2), and her systemic examination was unremarkable. Nonetheless, a 12-lead ECG done at our center revealed deeply inverted T-waves on leads V2 through V4 , which revealed a nonthrombogenic severe subocclusive (>95%) mid-LAD stenosis with a TIMI I flow . The lef1012 Young patients presenting with chest pain may often appear deceptively well with no apparent risk factors or hemodynamic instability10; however, clinicians should strive to recognize it, exclude life-threatening causes, and intervene timely. Owing to the increasing prevalence of CAD risk factors in younger individuals, the traditionally known protection offered by young age is slowly fading and an increased disease burden will be apparent in the near future.13With an estimated contribution of up to 6% of all acute coronary events, young adults (<45 years) are experiencing an escalating burden and mortality attributable to CAD globally.19 Despite of its known diagnostic and prognostic value, recognition of Wellens syndrome remains a challenge to practitioners all over the world.19 As demonstrated in this case, a definitive diagnosis was reached on the eighth day since onset of chest pain. Arguably, this was so due to various reasons: (1) absence of pathognomonic features for Wellens syndrome in an initial ECG, (2) absence of traditional risk factors for CAD in this case that probably resulted in lack of aggressive workup including serial ECGs, and (3) poor understanding and low index of suspicion for Wellens syndrome among cardiovascular physicians working in the initial consultant referral hospital.Wellens syndrome is an ECG manifestation characterized by a pattern of deeply inverted or biphasic T-waves in leads V2 to V3, signifying a critical stenosis of the LAD coronary artery.19 Nevertheless, presence of positive cardiac biomarkers portends a poorer prognosis and warrants urgent catheterization as it has been associated with a higher incidence of total LAD occlusion.6 Furthermore, like in this case, stress test should be avoided in individuals with Wellens syndrome as it is associated with an increase in myocardial demand, which may induce MI, LV failure, and subsequent sudden death.5 As this case met the criteria for type B Wellens syndrome, which is comparatively less common  than type A, it is worth mentioning that it is associated with higher fatality.14Cardiac enzymes may give false reassurance in patients with Wellens syndrome as they are frequently negative as it was in our case.The ability to accurately interpret abnormalities on ECG is a core competency for cardiovascular and emergency physicians that is central to the assessment of patients with potential cardiac ischemia. High-risk ECG patterns inform the diagnosis and management of patients with suspected acute coronary syndrome, but the failure to identify it or the impact of misinterpretation of the tracing is likely to result with a grievous consequence. Luckily, despite the delay in reaching the diagnosis, ECG performed at our institute revealed the typical Wellens syndrome features and was correctly interpreted by the attending cardiologist. Nonetheless, even after diagnosis realization coupled by ongoing chest pain, there was yet a further 3-day delay in taking the patient for revascularization . As a consequence of cumulative delays both in diagnosis and intervention, and notwithstanding successful revascularization, echocardiographic assessment done 1 month later revealed persistent ischemic features suggestive of a significant myocardial injury.16 Considering the presence of severe anemia coupled by absence of acute thrombotic evidence in coronary angiography, and assuming that there was a significant preexisting LAD disease, it is likely that a combination of such events triggered Wellens syndrome in the case presented.Last, anemia in the setting of CAD has an established adverse prognostic value. It is associated with new onset and recurrence of acute coronary syndrome as well as increased risk for mortality both short and long term.To conclude, Wellens syndrome is a relatively common yet underrecognized preinfarction entity with an ill-fated MI progressive potential if unintervened. In view of this, physicians should be well aware of this subtle yet warning ECG sign as prompt recognition and a timely aggressive intervention is of paramount importance to curb the attributable morbidity and mortality. Moreover, presence of angina particularly in healthy young patients with no apparent risk factors for CAD should raise an index of suspicion for Wellens syndrome.


Insulin is a hormone typically associated with pancreatic release and blood sugar regulation. The brain was long thought to be “insulin-independent,” but research has shown that insulin receptors (IR) are expressed on neurons, microglia and astrocytes, among other cells. The effects of insulin on cells within the central nervous system are varied, and can include both metabolic and non-metabolic functions. Emerging data suggests that insulin can improve neuronal survival or recovery after trauma or during neurodegenerative diseases. Further, data suggests a strong anti-inflammatory component of insulin, which may also play a role in both neurotrauma and neurodegeneration. As a result, administration of exogenous insulin, either via systemic or intranasal routes, is an increasing area of focus in research in neurotrauma and neurodegenerative disorders. This review will explore the literature to date on the role of insulin in neurotrauma and neurodegeneration, with a focus on traumatic brain injury (TBI), spinal cord injury (SCI), Alzheimer’s disease (AD) and Parkinson’s disease (PD). Insulin receptors (IR) are expressed by neurons and glia and mediate insulin signaling throughout the brain . While ide novo , the majInsulin signaling plays a role in global brain glucose metabolism  and cereIn order to achieve the positive cognitive and mood effects of insulin, without altering systemic blood glucose levels, alternative delivery methods have been explored. Evaluation of memory in rodent models demonstrated elevated hippocampal neurogenesis and BDNF production following direct infusion of insulin to the cerebral ventricles . IntranaIntranasal delivery of insulin allows insulin to bypass the saturable BBB insulin delivery system and reach the brain directly via the olfactory and trigeminal nerve pathways and distribution into the cerebrospinal fluid (CSF) . Thus, iReduced sensitivity or resistance to insulin actions, via downregulation or loss of IR or reduced activity of insulin signaling pathway, contributes to worsened outcome in several neurological conditions, further highlighting the importance of insulin in the CNS. Insulin resistance is observed in various instances of neurotrauma  and neurThe focus of this review is therefore to characterize the current literature on the role of insulin in CNS disorders, with a focus on traumatic brain injury (TBI), spinal cord injury (SCI), and neurodegenerative diseases, specifically Alzheimer’s disease (AD) and Parkinson’s disease (PD).In vivo studies demonstrated insulin’s cognitive effects by showing that inhibition of endogenous intrahippocampal insulin impaired spatial working memory in the spontaneous alternation behavior in a 4-arm plus-maze (SA) and acute administration of intrahippocampal insulin enhances SA performance , and insulin/IGF-1 hybrid receptors. Insulin signaling is modulated through the tyrosine phosphorylation of cellular substrates, including several IR substrates (IRS) , as wellformance . To elucognition . Additioognition . Insulinognition .Insulin signaling is fundamental for proper functioning of neurons and memory formation and storage. There is an increase in IRs in synapses of the hippocampus following a short-term memory task . Insulinex vivo mouse model of diabetes, where cortical slices from diabetic BKS-db/db mice demonstrated no change in phosphorylation of AKT when treated with insulin. In contrast, cortical slices from controls did show a dose-dependent increase in AKT phosphorylation. These results indicate that chronic insulin stimulation results in decreased acute insulin-stimulated AKT activation; this is a situation that should be kept in mind when designing long-term therapeutic strategies utilizing insulin.Primary rat cortical neurons have been used to mimic insulin resistance . Corticain vitro model of excitotoxicity  pathways. Full activation of the IIS pathway in human cultured astrocytes was identified using western blot analysis for expression of IRβ, IGF1Rβ, IRS1, IRS2, pAKT, Total AKT, p44/42, and MAPK following insulin treatment (1 μM recombinant human insulin) .in vitro to respond to insulin at low doses (1 nM) with increased production of IL-6 and IL-8, pro-inflammatory cytokines. However, this effect dissipates at higher doses (100 nM), suggesting that insulin plays a role in astrocyte inflammatory response as well as glycogen storage  protects against 24 h MPP + (500 μM) stimulation-induced toxicity, as demonstrated by significantly decreased LDH and nitric oxide (NO) media release and iNOS and COX2 expression when compared to MPP + stimulated only cells  from mice fed a high fat diet (HFD), consisting of 60% energy from fats for 7 days, have been used as a model of insulin resistance to study effects on inflammation. Isolated TEPMs, from mice on HFD, exposed to a 30 min insulin treatment (100 nM) displayed a decrease in AKT phosphorylation and significantly lower expression of IR compared to TEPMs isolated from mice fed normal diets. TEPMs exposed to high-dose (100 nM) insulin for 48 h showed reduced AKT phosphorylation and decreased IR expression following a 30 min re-stimulation of insulin . FurtherTreatment with insulin also prevented LPS induced pro-inflammatory responses in primary mouse peritoneal macrophage . Cells sWith these cellular effects of insulin mind, we now move to consideration of insulin within different trauma and neurodegenerative conditions, to further explore the role of insulin in the CNS.Traumatic brain injury is a major cause of death and disability in the United States. There are two primary injury types of TBI, focal and diffuse, although many injuries can present with both components. Focal injuries are direct area injuries resulting from collision or penetrative forces acting upon the skull and commonly present with contusions and subdural hematomas . DiffuseCo-morbidities are not uncommon in TBI; head injuries greatly increase the risk of neurodegenerative diseases such as AD and chronic traumatic encephalopathy . AdditioTraumatic brain injury is a pathologically heterogenous disease, which can result in a variety of cognitive deficits depending on the location, type, and severity of damage . Up to 1Post TBI cognitive deficits can also result from cellular and metabolic dysfunction after injury, including inflammation, insulin resistance, and decreased cerebral glucose uptake. As a direct result of the injury, necrotic neuronal cell death can occur  and an iThese cellular alterations result in a fluctuation of glucose uptake and metabolism that have often been referred to as the neurometabolic cascade of concussion , though These alterations are observed in clinical work as well. Many patients demonstrate hyperglycemia, which has been found to be a predictor of poor neurological outcome . AdditioHyperglycemia may be stress-induced, which is caused by activation of both the hypothalamic-pituitary-adrenal axis and sympathetic autonomic nervous system after TBI . Both sy18F]-FDG uptake profile after moderate controlled cortical impact injury (CCI) is altered by injury and dependent on the cellular composition of the region of interest (18fluorodeoxyglucose (FDG) positron emission tomography (PET).Our lab has shown that a “milder,” diffuse lateral fluid percussion injury in rats resulted in global cerebral hypometabolism that persisted from 3 h to 9 days post injury . We haveinterest . These dSeveral clinical trials have aimed to treat hyperglycemia after TBI by glucose control via IV administration of insulin , but resOur previous work demonstrated that after a moderate TBI in rats, intranasal insulin administered within 4 h after injury significantly increased glucose uptake in the hippocampus, improved cognitive function, and reduced inflammation . In thisin vitro, mice fed HFDs for 12 weeks demonstrated elevated levels of pro-inflammatory cytokine gene expression the hippocampus that were reduced by hippocampal infusion of insulin (2.64 μL of Humulin insulin per day at 4 mU/μL)  . With th4 mU/μL) , althoug4 mU/μL) . The litWithin the United States alone, there are approximately 12,000 new cases of SCI each year . This tyAfter SCI, peripheral blood glucose is significantly altered. Female mice exposed to a thoracic contusion injury demonstrated elevations in blood glucose levels from 4 h through at least 2 weeks post-injury, accompanied by significant reductions in peripheral insulin levels . These cIn mice exposed to SCI, insulin levels are reduced from 7 to 28 days post-injury (the longest time point studied) . ImpairmPerhaps unsurprisingly, SCI is associated with an increase in occurrence of Type II diabetes and peripheral insulin resistance . In a stAlthough CNS insulin resistance has not yet been evaluated after SCI, this type of injury has been found to significantly alter glucose uptake in injured spinal cord tissue; this may be due to changes in availability of peripheral glucose or alterations in ability of damaged tissue to take up glucose. Autoradiography examination of monkey spinal cord demonstrated that glucose metabolism was acutely increased for 1 h after injury , followeIn addition, we demonstrated that these changes in glucose uptake were dependent on subject age. Increasing the age of rats from 3 months  to 12 moChronically, autoradiography studies have shown that glucose uptake and metabolism is depressed in the moderately injured rodent spinal cord . GlucoseThe effect of SCI on IR expression in the brain or spinal cord has not yet been evaluated. However, GLUT3 and 4, both of which show insulin dependence, have been shown to be elevated at both the protein and gene level 2 weeks after injury in both young and aged populations . Previouin vitro or injections of streptozotocin in vivo have been shown to increase pro-inflammatory markers and neuronal damage  . CombiniIn a rabbit model of ischemic SCI, a single dose of insulin (0.5 IU/kg) peripherally 30 min prior to injury significantly improved motor function and neuronal survival at 7 days post-injury . This peThese data demonstrate that insulin delivery after SCI has potential, although a risk of hypoglycemia suggests treatment should proceed carefully. As in TBI, central administration of insulin may prove to be a useful alternative. However, essential basic biology into IR expression and effects of SCI are needed to further explore therapeutic options.Insulin signaling impairment and insulin resistance have been observed in neurodegenerative diseases such as AD  and PD  Figure . DiabeteAlzheimer’s Disease is the most common neurodegenerative disease and form of dementia. It is characterized by cognitive and functional decline over time . AmyloidStudies have shown that AD presents with its own neurometabolic response prompting some researchers to refer to AD as “type 3 diabetes” . This teA major branch of the insulin signaling pathway, the PI3K/AKT pathway, is downregulated in AD and this change may be a major contributor to insulin resistance . IRS1 isParkinson’s disease is the second most common neurodegenerative disorder and is characterized by decreased movement, walking instability, tremors, and associated dementia . Many ofInsulin plays a role in dopamine release in the brain; when present insulin enhances dopamine uptake via the PI3K pathway  and dopaIntranasal insulin has been explored as a treatment for AD and PD in both animal and clinical studies . It has In the six OHDA rat model of PD, both a daily high dose (12IU) for 2 weeks and a daily low dose (3IU) for 4 weeks of intranasal insulin alleviated motor deficits observed in the model in overall locomotor activity and in a variety of motor behavioral tests . Pang etIn 2008, Dr. William Frey II’s group at the University of Minnesota reported that intranasal insulin was effective in improving memory for patients suffering from AD disease, and the patients suffered no adverse effects of decreased blood sugar . ClinicaAdditionally, in studies comparing memory impaired/AD participants with and without a risk factor for late onset AD, the ApoE e4 allele, insulin effects varied based on the genotype , 2003. IIn a recent PD clinical trial, intranasal insulin treatment caused increased verbal fluency (FAS) in participants that received treatment, while the placebo group had decreased FAS scores indicating that there was improvement in cognition in the treated group. Participants treated with insulin also had lower disability scores compared to the placebo group, demonstrating improvement in motor function and overall performance with treatment .Intranasal insulin has been shown to produce beneficial effects in healthy participants. It can enhance feelings of self-confidence and well-being, thus improving overall mood, as well as aid in word recollection in a word list learning task . ImproveAnother method of insulin delivery, IV administration, in AD patients has showed improved memory performance , but lonIn vitro and in vivo studies demonstrate insulin plays a significant role in cellular metabolism in all brain cells and disruption of insulin signaling, from insulin resistance, can contribute to dysfunction and pathological conditions as observed in AD and PD. Clinical trials using insulin, particularly intranasal insulin, show promise in patients with AD and PD, as treatment leads to restoration of insulin signaling and amelioration of cognitive and motor deficits. Our lab has shown the therapeutic effects of intranasal insulin in an experimental model of TBI, that improves memory, increase glucose uptake, and decreases neuroinflammation and hippocampal lesion volume. This review highlights the therapeutic value of insulin in the context of neurotrauma and several neurodegenerative disorder and describes the cellular effects of insulin resistance on multiple brain cells. Further research is warranted to identify dosing and schedule of insulin treatment to improve outcomes for patients suffering from neurological disease and trauma, and to understand the role insulin resistance plays in the development and progression of brain disorders.The therapeutic value of insulin and the deleterious effect of insulin resistance on neuronal and glia cells, as it related to neurotrauma and neurodegeneration, has only recently been explored. in vitro section. DA researched, wrote neurodegeneration section, and designed figures. FB, DA, and NH researched and wrote traumatic brain injury section. FB and MS wrote introduction. All authors contributed to the article and approved the submitted version.KB designed the overview of the manuscript and wrote spinal cord injury section. MS researched and wrote The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The retinal ganglion cells (RGCs) are the output cells of the retina into the brain. In mammals, these cells are not able to regenerate their axons after optic nerve injury, leaving the patients with optic neuropathies with permanent visual loss. An effective RGCs-directed therapy could provide a beneficial effect to prevent the progression of the disease. Axonal injury leads to the functional loss of RGCs and subsequently induces neuronal death, and axonal regeneration would be essential to restore the neuronal connectivity, and to reestablish the function of the visual system. The manipulation of several intrinsic and extrinsic factors has been proposed in order to stimulate axonal regeneration and functional repairing of axonal connections in the visual pathway. However, there is a missing point in the process since, until now, there is no therapeutic strategy directed to promote axonal regeneration of RGCs as a therapeutic approach for optic neuropathies. The retina is part of the central nervous system (CNS) and is constituted by neurons, glial cells and blood vessels . The neuOptic neuropathies comprise a group of ocular diseases, like glaucoma (the most common), anterior ischemic optic neuropathy and retinal ischemia, in which RGCs are the main affected cells . BlindneDuring development, RGCs extend their axons to synapse in target areas of the brain (reviewed in ). After 2+) increases in amacrine cell processes upon optic nerve injury and is transferred to RGCs via vesicular release [2+ improves cell survival and axon regeneration [2+ levels is responsible for the loss of axonal growth.cAMP plays an important role in neuronal survival and axon growth and guidance . For exa release . The cheneration , raisingOther transcription factors have been studied for their role in axon growth and regeneration (reviewed in ). The tuGlial scar and myelin that compose the environment of optic nerve particularly at the site of injury inhibit the axonal regeneration (reviewed in ). SemaphThe failure to regenerate has also been attributed to an environment poor in growth-promoting trophic factors. In fact, the importance of trophic factors in promoting viability and axonal regeneration of RGCs has long been recognized . A greatThe survival of RGCs is increased by co-administration of BDNF and CNTF soon after optic nerve injury . MoreoveStudying the mechanisms of glaucomatous damage has been a great opportunity to unravel the signaling pathways involved in RGC axonal degeneration and growth. Elevated IOP is the main risk factor of glaucoma and, together with other factors, it has been implicated in RGC degeneration and death . SeveralRecently, it was reported that intravitreal injections of BDNF leads to an increase in the levels of synaptic proteins between RGCs and bipolar cells in the IPL, meaning that this could have a beneficial effect in the function of RGCs .Several therapeutic strategies have been proposed in order to protect RGCs and restore visual function .Neurotrophic factors are a family of growth factors that regulate the survival, development and differentiation of neurons. Neurotrophic factors generally include the neurotrophin family: NGF, BDNF, neurotrophin-3 (NT-3) and neurotrophin-4/5 (NT-4/5); the glial cell-line derived neurotrophic factor (GDNF) family: GDNF, neurturin (NRTN), artemin (ARTN), and persephin (PSPN); and CNTF ,81. It wNGF is an important growth factor affecting the survival of nerve cells and their deprivation can lead to apoptosis ,91. NGF BDNF is widely expressed throughout the CNS. RGCs express BDNF and its high affinity receptor TrkB . As mentGDNF is secreted by glial cells and binds to the GDNF-α receptor and to the receptor tyrosine kinase in RGCs . GDNF prCNTF belongs to the interleukin-6 (IL-6) family of cytokines, binds to CNTF receptors (CNTFR) and exerts robust neuroprotection in neurons ,111. In Other trophic factors have been described to promote RGCs survival. Pigment epithelium derived factor (PEDF) reduces RGC loss in a mouse model of glaucoma  and insuDespite being the major excitatory neurotransmitter in the retina and involved in the vertical pathway of information , excessiThe presence of alpha-adrenergic receptors in the RGCs has been demonstrated . AdditioCalcium channel blockers may protect RGCs by preventing cell death mediated by calcium influx secondary to NMDA receptor overactivation and local ischemia ,150. DifOxidative stress occurs when concentrations of reactive oxygen species (ROS) rise above physiological range, and it has been indicated as a potential cause of glaucomatous neurodegeneration . Thus, iGlutathione (GSH) is decreased in glaucoma patients, suggesting a general compromise of the antioxidative defense . The treThe levels of nitric oxide (NO) are increased in experimental glaucoma, and evidence shows that NO can result in RGCs degeneration ,174,175.1, A2A, A2B and A3. Adenosine acting on adenosine A1 receptor (A1R) protects cultured retinal neurons from NMDA-induced cell death [1R could be a good therapeutic strategy. Indeed, the activation of A1R is neuroprotective against injury induced by ischemia-reperfusion [1R, increases RGCs survival mediating the trophic effect of IL-6 [3 receptor (A3R) has also been evaluated as a therapeutic target [3R [3R promotes RGCs neurite outgrowth and neurite regeneration in an animal model of ONC [3R was also demonstrated to afford protection to the retina from excitotoxic-induced cell death, retinal ischemia-reperfusion injury and damage induced by partial optic nerve transection [Adenosine can exert both neuroprotective and neurodegenerative actions acting through four types of receptors: All death  by blockll death , suggesterfusion , and N derived from bone marrow, adipose tissues and dental pulp .Various methods to assess the ability of RGCs to survive and integrate with host tissue have been proposed ,207. TraMSCs have been widely demonstrated to afford neuroprotective, immunomodulatory and antioxidant properties, making them a promising strategy for the treatment of neurodegenerative diseases. These cells secrete neurotrophic factors like NGF accelerating the survival of neural cells . The proThe term neuroinflammation comprises a number of events that affects the CNS. In other words, every time the CNS is faced with infectious agents, traumatic injuries or other unknown elements that might cause a disruption of its homeostasis, it will protect itself by the initiation of inflammatory signaling cascades in order to eliminate the pathological factor . AlthougMicroglial cells have long been recognized as crucial players in the maintenance of retinal homeostasis. During development, microglial cells are involved in synaptic pruning and in retinal wiring  and thro2A receptor (A2AR) may propel microglia deleterious response overtime [2AR has been described to control microglia reactivity. Its expression increases in microglia in models of glaucoma [2AR antagonists were shown to confer protection to retinal neurons, including RGCs, through the control of microglia reactivity [2AR antagonist with good oral bioavailability, confers protection to the retina, including RGCs, through the control of microglia-mediated neuroinflammation [2AR antagonists was further confirmed as a strategy for the human retina [2AR antagonist was able to reduce microglia alterations and the production of ROS, suggesting that microglia-mediated inflammation in the human retina also involves A2AR [The pivotal role of microglia in RGCs degeneration has been mostly explored in glaucoma. Historically, reactive microglial cells have been associated with human glaucomatous ONH lesion, mainly by their spatial distribution along the damaged fibers and expression of activation markers as well as pro-inflammatory mediators ,225. Indovertime . The A2Aglaucoma ,79, and activity ,79,228. activity ,234,235.n retina . By usinves A2AR . The neuves A2AR . In suchves A2AR .The secretion of TNF by microglial cells was shown to contribute to RGC degeneration as its receptor is highly increased in glaucoma in RGCs, astrocytes, microglia and Müller cells, triggering a cascade of events that culminates in RGC demise ,237. IndRecently it has been shown that chronic OHT promotes the expression of P2X7 receptor in the retina leading to the activation of NLRP3  inflammasome . The actMicroglial cells also modulate retinal cell function by expressing complement molecules. In fact, in human and experimental glaucoma the expression of complement factors is increased in conditions of elevated IOP ,242,243.In the model of ischemia-reperfusion, treatment with minocycline, which decreases microglia reactivity, was able to protect RGCs . MoreoveMüller cells are the main glial cells in the retina. In addition to structural support, among other functions, Müller cells are involved in metabolism, phagocytosis of neuronal debris and in the release of trophic factors. These cells can enhance the survival of RGCs ,251,252.Müller cells are crucial to protect neurons against toxic molecules . They caMüller cells are also involved in the regulation of glycogen and glucose metabolism and during metabolic stress can provide lactate to retinal neurons . For insMüller cells secrete a great number of factors in response to injury that can protect retinal neurons. Müller cells are known to synthesize neurotrophins and growth factors that can increase RGC survival ,252,254,Furthermore, Müller glial cells can release several inflammatory factors and cytokines , and somA therapeutic strategy to optic neuropathies should protect RGCs from death but should also manipulate axonal regeneration in order to repair the visual function that was lost due to the disease. However, there is still no effective therapy for optic neuropathies. Innovative study designs and integrating therapeutic testing with biomarkers have advanced several neuroprotective and neuroenhancement compounds to clinical trials. Numerous neuroprotection strategies have been investigated for optic neuropathies, including peripheral nerve grafting, electrical stimulation, and in agreement with their well-known role in maintaining neuronal homeostasis, neurotrophic factors have been proposed as a novel therapy. However, the outcomes of the completed clinical trials were not completely satisfactory, presenting only partial or no expected effects ,280,281.ClinicalTrials.gov Identifier: NCT02862938) and in phase 1 for ischemic optic neuropathy . For glaucoma, other therapies have been proposed such as the use of recombinant human NGF (rhNGF) . In this phase 1 clinical trial the safety and tolerability of an 8-week treatment with 180 μg/mL of rhNGF eye drop solution will be determined. Additionally, the study wants to assess the changes in best corrected distance visual acuity (BCDVA), visual field, electroretinography (ERG) and structural changes in GCL and NFL thickness measured by optical coherence tomography (OCT) at 1, 4 and 8 weeks of therapy, and at 4 and 24 weeks after therapy cessation. In another clinical trial the safety of treatment with single and multiple ascending doses of rhNGF (0.5–180 µg/mL) was tested in healthy patients , and the results demonstrated that rhNGF eye drops were well tolerated by the patients [There are several drugs in clinical trials that are currently being developed focused on RGC neuroprotection . In the patients .ClinicalTrials.gov Identifier: NCT00693485). Moreover, cytidine-5′-diphosphocholine (citicoline) is also in a phase 4 clinical trial for glaucoma . Citicoline is an endogenous molecule that has a role in the biosynthesis of phospholipids of cell membranes and increases the levels of neurotransmitters, like acetylcholine, in the CNS [ClinicalTrials.gov Identifier: NCT00141882 and NCT00168350). However, the drug did not show significant efficacy in preserving visual function in glaucoma patients [The only modifiable risk factor for glaucoma development is elevated IOP. Brimonidine is a non-selective α2-adrenergic receptor agonist and is currently used as a treatment option in glaucoma to lower IOP . Preclin the CNS . The neu the CNS ,287. Int the CNS , RGC fun the CNS . That wapatients .ClinicalTrials.gov Identifier: NCT03851562). Prostaglandin E1 is a potent vasodilator of the microcirculation [ClinicalTrials.gov Identifier: NCT02377271). The last drug-based therapy for ischemic optic neuropathy, the retrobulbar injection of triamcinolone acetonide to halt the progression of the visual acuity and visual field loss in patients improving their chances of avoiding blindness, is in phase 3 clinical trial . In preclinical studies, besides the neuroprotective effects to RGCs conferred by triamcinolone acetonide, it was demonstrated that this drug also decreases the activation of retinal microglia [ClinicalTrials.gov Identifier: NCT03715881). In the same clinical trial, another aim is to assess the potential retinal neuroprotective effect of prednisolone. Moreover, methylprednisolone is also in phase 3 clinical trial . Preclinical studies demonstrated that methylprednisolone inhibits the apoptosis of RGCs after ONC, probably through an up-regulation of Bcl-2 expression and a down-regulation of Bax expression [ClinicalTrials.gov Identifier: NCT03046693) in order to assess the function of RGCs by pattern ERG, thickness of GCL and visual field test.Moreover, prostaglandin E1  administered by intravenous infusion, is very recently in phase 2 clinical trial . The results of the phase 2 clinical trial  are already available. Patients showed an improvement in visual function after the treatment [ClinicalTrials.gov Identifier: NCT01975324).RPh201 is a drug extracted from a botanical source and it has been produced by Regenera Pharma. RPh201 started recently the phase 3 clinical trial for non-arteritic ischemic optic neuropathy  in order to halt the progression of visual acuity and visual field loss due to the disease. The thickness of GCL increased after the treatment with bevacizumab in diabetic macular edema [ClinicalTrials.gov Identifier: NCT00432393).Anti-VEGF antibodies  are used for the treatment of macular edema and neovascular age-related macular degeneration. However, they have also been tested for neuroprotection in optic neuropathies, and they are in three different clinical trials for non-arteritic anterior ischemic optic neuropathy  assessing the effect of erythropoietin (EPO) demonstrated an improvement in visual function [ClinicalTrials.gov Identifier: NCT02376881). EPO is currently in phase 3 clinical trial for traumatic optic neuropathy .A phase 1 and 2 clinical trial . The beneficial effects of idebenone are due to its antioxidant properties and its ability to act as an electron carrier in the mitochondrial respiratory chain, thus resulting in the restoration of cellular energy (ATP) generation and contributing to the recovery of visual function in patients (reviewed in [®) is the first, and currently the only disease-specific treatment for Leber’s hereditary optic neuropathy and the only approved for optic neuropathies aiming RGCs neuroprotection. Moreover, cyclosporine is also in a phase 2 clinical trial for Leber’s hereditary optic neuropathy , due to protective properties against ischemic injury-mediated mitochondrial dysfunction in RGCs [Leber’s hereditary optic neuropathy is an inherited optic neuropathy characterized by mitochondrial dysfunction that leads to vision loss due to RGCs loss . Idebenoiewed in ). That w in RGCs .ClinicalTrials.gov Identifier: NCT02638714) on restoring function in damaged optic nerves using autologous purified populations of bone-marrow derived stem cells in optic neuropathy. The intravitreal injection of MSCs  aims to evaluate if the treatment may reduce the progression of axonal degeneration caused by non-arteritic ischemic optic neuropathy, but this clinical trial is focused in the evaluation of the safety of cell therapy as a new treatment for these patients.Currently, there are two clinical trials involving stem-cell based therapies targeting RGCs . One triThe complexity of the CNS is due to the great number of specialized neuronal types and subtypes that give rise to a complex connectome . HoweverThe major requirement to properly characterize and classify RGCs is to distinguish selectively each specific subtype. Thus, recently, a number of molecular subtype-specific markers have been described to further classify different subtypes of RGCs . SeveralAnother molecular marker, Tbr2, identifies RGCs that are hardwired during developmental stages and they could be precursors of ipRGCs . Other sMoreover, mouse transgenic lines have been also used to label and identify specific subsets of RGCs . For insIn order to understand the pathophysiology of neurodegenerative diseases in which the RGCs death is implicated, it is important to analyze the response of these RGCs subtypes individually rather than studying them as a single entity. RGCs are susceptible to various injuries in a type-specific manner. Thereby, their type-specific vulnerability has been extensively studied . The ideIt has been shown that non-image forming ipRGCs exhibited a preferential survival following injury compared to image forming RGCs. This fact was observed in different injury and disease models, demonstrating the resilience to damage of this subtype of RGCs . ipRGCs All these studies clearly imply that RGCs respond in a subtype specific manner to injury. Moreover, each subtype of RGCs can have a unique gene expression pattern ,329, thiThe main goal in finding new therapeutic strategies for optic neuropathies is to preserve the function of RGCs in order to maintain visual pathways. Therefore, besides the neuroprotection of RGCs and axon regeneration, the re-integration of RGC axons into the appropriate visual circuity is also important. Despite that several therapeutic strategies have demonstrated promising results in this field, there are significant issues affecting their translation to clinical practice.Much work has been done in order to identify the inhibitors of axonal growth in the CNS as well as to isolate neurotrophic factors, with the hope that one day these factors could be applied to protect and regenerate the optic nerve. From what was described above, it seems that we are getting closer to a therapeutic strategy focused on RGC neuroprotection for optic neuropathies. However, we can discuss the example of memantine that, despite the convincing neuroprotective effects in animal models of glaucoma, in clinical trials the drug did not reveal significant effects in preventing the progression of visual field loss in patients with glaucoma .Several issues contributing to the lack of success of drugs in clinical trials could be suggested, but in the case of glaucoma, the lack of an animal model that fully mimics the human disease is an important factor that adds to this failure . AnotherBesides the protection of RGCs from death and degeneration, one of the goals in RGCs regeneration therapies should be to allow the reintegration of the regenerating axons into visual circuity reaching the appropriate brain targets. However, there are few studies that focus on this issue . MoreoveIt is fundamental a better characterization of the beneficial effects of drugs in the preclinical phase, meaning that the observation of the loss of RGCs is not enough as it is not enough to observe potential regenerative events of the axons of RGCs. It is essential to clearly and more deeply evaluate the beneficial effects of a specific new drug also in visual function.The research in the field of neuroprotection in glaucoma has been difficult, but new animal models of disease and techniques will help to bridge the gap between preclinical and clinical studies, with clear beneficial outcomes in the forthcoming years. Many of the approaches outlined in this review are applicable not only to RGC neuroprotection in glaucoma but also to other pathologies of the optic nerve and retina. Gene therapy may have also a therapeutic potential especially for Leber’s hereditary optic neuropathy, an optic neuropathy caused by mitochondrial mutation G11778A in NADH dehydrogenase subunit 4 (ND4) gene . It was


Currently, there are no disease-modifying treatments for Alzheimer’s disease (AD) or any other dementia subtype. The renaissance in psychedelic research in recent years, in particular studies involving psilocybin and lysergic acid diethylamide (LSD), coupled with anecdotal reports of cognitive benefits from micro-dosing, suggests that they may have a therapeutic role in a range of psychiatric and neurological conditions due to their potential to stimulate neurogenesis, provoke neuroplastic changes and reduce neuroinflammation. This inevitably makes them interesting candidates for therapeutics in dementia. This mini-review will look at the basic science and current clinical evidence for the role of psychedelics in treating dementia, especially early AD, with a particular focus on micro-dosing of the classical psychedelics LSD and psilocybin. Globally an estimated 50 million people have a diagnosis of dementia and population prevalence continues to increase  that, in accumulation, are associated with a variety of pathological processes including microtubular damage, axonal transport disruption and, ultimately, cell death. The hippocampus, a key structure in the ability to learn and retain information and a site for neurogenesis, is particularly vulnerable to AD pathology and one of the earliest parts of the brain to be affected by the disease  agonists, in psychiatric and neurological disorders Nichols, . The 5HTvia the 5HT2A-R in regions such as the hippocampus could theoretically help protect this and other brain structures and may, therefore, hold potential for treating AD.This mini-review will explore the role of classical psychedelics psilocybin and Lysergic acid diethylamide (LSD) in treating AD with a focus on sub-perceptual- or “micro”- dosing. Promoting neuroplasticity and neurogenesis per se  rather than reduced attentional ability n = 20 and n = 24, respectively), both using a within-subjects design, found no effects either positive or negative on the cognitive function of healthy volunteers at different sub-perceptual doses  who had not taken LSD for at least the past 5 years found no difference in the number of adverse events (including cognitive impairment) between those taking placebo, 5, 10, or 20 μg doses every 5 days for 28 days  enhances both prospective and retrospective learning with lesser effects at low-dose , led to clinical response or remission in 14 participants sustained at assessment 5 weeks later. This effect persisted at 6 months follow up despite no further treatment  expression in the neocortex but appears to consistently inhibit the same process in the hippocampus  and is completely reversed by 5 days of abstinence  within neuronal networks are important for communication between brain regions, particularly those involving attention and memory  has been shown to attenuate amygdala reactivity to emotional stimuli (Kraehenmann et al., Studies suggest that at least some of the antidepressant effect from psilocybin may be mediated These imaging studies reveal the potent reorganization of dysfunctional brain networks in affective and anxiety states. Such changes may also yield improvements in cognition, mood, and behavior by ameliorating dysfunctional circuits in cognitive impairment and dementia.All known genetic and environmental risk factors for AD are associated with increased inflammation, suggesting that reducing inflammation could be a target for preventing AD (Jones and Kounatidis, via anti-apoptotic pathways (Shahidi et al., via the upregulation of neuroprotective Sirtuin 1 expression (Fanibunda et al., In a rodent model of AD induced by chronic intracerebral inoculation of streptozotocin, 5HT1A- and 5HT2A-R agonists had a significant independent and synergistic neuroprotective effect in hippocampal neurons at 35 days After decades of repeated failure of treatments for dementia, there is an urgent need to develop new treatments for AD. The potential for psychedelic compounds to influence and enhance functional neuronal connectivity, stimulate neurogenesis, restore brain plasticity, reduce inflammation and enhance cognition provides a new therapeutic target and compelling argument for further investigation of the potential for psychedelics as a disease-modifying compound in conditions where currently none exists.Animal models testing the neurobiological effects of psychedelic compounds have demonstrated hippocampal neurogenesis at lower doses and suppression at higher doses and potent neuroprotective properties. Studies in people suffering from depression and anxiety disorders have demonstrated lasting neuroplastic changes following just one or two large doses. This suggests a potential role for both sub-perceptual “micro”- and psychedelic-doses as a strategy for neuroprotection and cognitive enhancement in prodromal AD. For cognitive enhancement, the ideal dose and frequency have yet to be determined however the rapid desensitization of 5HT2A receptors by both psilocybin and LSD suggests that daily dosing is unlikely to be the optimal strategy.Despite anecdotal evidence of widespread recreational use of micro-dosing for cognitive enhancement, robust scientific studies of the cognitive effects of micro-dosing in humans have so far been limited to acute changes in very small studies in cognitively normal individuals with no reports of persistent cognitive changes, either positive or negative, at psychedelic doses. Studies looking at both micro-dosing and psychedelic doses, longer-term, in cognitively impaired individuals are lacking and urgently needed.SV came up with the idea for the review. Both SV and AO’K were involved in a review of the literature and writing and editing the final manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Additionally, the cardiovascular system is quite sensitive in responding to changes in mechanical stress from internal and external environments. Several mechanotransduction pathways are involved in regulating the physiological and pathophysiological status of cardiomyocytes. Typically, the extracellular matrix generates a stress-loading gradient, which can be sensed by sensors located in cellular membranes, including biophysical and biochemical sensors. In subsequent stages, stress stimulation would regulate the transcription of mitochondrial related genes through intracellular transduction pathways. Emerging evidence reveals that mechanotransduction pathways have greatly impacted the regulation of mitochondrial homeostasis. Excessive mechanical stress loading contributes to impairing mitochondrial function, leading to cardiac disorder. Therefore, the concept of restoring mitochondrial function by shutting down the excessive mechanotransduction pathways is a promising therapeutic strategy for cardiovascular diseases. Recently, viral and non-viral protocols have shown potentials in application of gene therapy. This review examines the biological process of mechanotransduction pathways in regulating mitochondrial function in response to mechanical stress during the development of cardiomyopathy and heart failure. We also summarize gene therapy delivery protocols to explore treatments based on mechanical stress–induced mitochondrial dysfunction, to provide new integrative insights into cardiovascular diseases.Mitochondria are one of the most important organelles in cardiomyocytes. Mitochondrial homeostasis is necessary for the maintenance of normal heart function. Mitochondria perform four major biological processes in cardiomyocytes: mitochondrial dynamics, metabolic regulation, Ca The heart is an electromechanical organ that needs to beat thousands of times a day to provide enough blood supplement to the body  specification of cardiac progenitor cells, (2) formation of the linear heart tube, (3) cardiac looping, and (4) formation of the cardiac valve to form a mature beating heart. Normal mechanical stimulation is essential to maintain the normal physiological processes of CMs, including proliferation, differentiation, and maturation. Generally, there are three types of mechanical loading approaches to CMs, including shear stress, cyclic strain, and static stretching. Shear stress is generated by friction at the interface between the blood and the endocardium in the same direction as the blood flow  plays a repressive role under these conditions , through the Wnt/activin/nodal pathway Pandur, , Gata-4 PKP2 (encoding plakophilin 2), DSG2 (encoding desmoglein 2), DSC2 (encoding desmocollin 2), JUP (junction plakoglobin), and DSP (desmoplakin) for desmosome formation and CDH2  and CTNNA3 (encoding catenin-α3) for adherens junction formation  activation, and increased ROS production /endoplasmic reticulum (ER) calcium ATPase 2 (SERCA2)  and cytokine expression levels contribute to the changes in ECM stiffness observed in healthy and damaged myocardium tissues . MQC is a complex process that includes three main levels. The first level prevents mitochondrial damage and maintains mitochondrial stability through proteasome activation. The second level maintains the number and shape of mitochondria through mitochondrial biogenesis, fission, and fusion. The third level selectively removes damaged mitochondria and is called mitophagy  is a critical factor in this process. PGC1 activates peroxisome proliferator-activated receptor, which leads to the gain of mitochondrial DNA (mtDNA) content, and promotes the transcription of mitochondrial uncoupling protein 1 (UCP1). This helps CMs adapt to the post-natal environment with a mature ETC utilizing fatty acid. PGC-1α expression is regulated by YAP/TAZ, and PGC-1α depletion results in lethal cardiomyopathy. Moreover, PCG-1 interacts with Mfn1/Mfn2 and Drp1. TFAM is another essential factor for mitochondrial biogenesis and is required for mtDNA transcription and mitochondrial self-replication. Zhang and Li demonstrated that the Nkx2.5-driven Tfam knockout leads to total mitochondrial loss in CMs  is the main mechanism of maintaining homeostasis. The UPS consists of ubiquitin, ubiquitin initiation enzyme, the 26S proteasome, and deubiquitination enzyme. Ubiquitin initiation enzyme is responsible for activating ubiquitin and target protein binding to form a target protein polyubiquitin chain or to ubiquitinate the degraded protein. Then, ubiquitinated proteins are transported to 26S proteasomes, where they are recognized and degraded  in the mitochondrial inner membrane. The proteasome complex can recognize misfolded transmembrane protein polypeptide chains and degrade unassembled mitochondrial complex subunits and transmembrane segments . PINK1 recruits phosphorylated Parkin through its silk/threonine kinase activity and transfers Parkin from the cytoplasm to the mitochondria. Parkin has E3 ubiquitin ligase activity and can mediate the ubiquitination of mitochondrial outer membrane proteins including HKI, VDAC1, and MFN1/2, to initiate mitophagy . DRP1 is highly expressed in heart tissues and can be upregulated under stress. Additionally, post-transcriptional SUMOylation and phosphorylation modulate DRP1 function. Mitochondrial fusion involves the joining of normal and damaged mitochondria to replace the materials in damaged mitochondria. This is conducive to material circulation and energy transmission and can protect cells. Mitochondrial fusion is mainly mediated by mitochondrial fusion proteins, including MFN1 and MFN2, which serve the outer mitochondrial membrane, and optic atrophy 1 (OPA1), which serves the intermitochondrial membrane. MFN1 and MFN2 have amino-terminal conserved GTPase domains and carboxy-terminal coiled helix structures and can bind to the mitochondrial outer membrane and form transhomologous or heterologous oligomeric complexes simultaneously, bringing two mitochondria into close proximity for fusion  is essential to maintain the electrochemical proton gradient across the mitochondrial inner membrane and should always remain 150–180 mV negative to the cytosol. ATP produced by mitochondria in myocardial tissue is mainly responsible for myocardial contraction, cellular excitability, and maintenance of calcium homeostasis. McCully et al. found that under myocardial ischemia and other pathological conditions, mitochondrial respiratory chain function is decreased, and ATP synthesis is impaired . The electrophysiological characterization and molecular identity of MCU are partially determined. MCU is an inward rectifying channel that is highly Ca2+-selective , SLC25A23 (solute carrier 25A23), and MCUR1  are important regulators of Ca2+ uptake , and the Na+/Ca2+ exchanger (NCLX)  (Guo et al., Recently, many reports have focused on the development of therapeutic compounds and drugs targeting mechanotransduction pathways to restore mitochondrial function in heart disease. For example, apigenin inhibits ROS generation, the loss of mitochondrial membrane potential, and apoptosis through PI3K/AKT signals and mitochondrial Notch1/HES1 signals, protecting H9C2 and rat heart cells from IR injury (Hu et al., In the last 10 years, ncRNAs have been studied worldwide, and several critical ncRNAs with essential biological functions have been identified. Antisense oligonucleotide therapy targeting strategies to modulate gene splicing have been used for ncRNA-based gene therapy. While lncRNA mimics have been used to express specific oligonucleotides (Lee J. et al., CRISPR/Cas9 has rapidly become one of the most popular and important approaches for genome editing because of its simplicity and adaptability. CRISPR/Cas9 and iPSC technology were recently applied to characterize Barth syndrome with mitochondrial abnormalities. The administration of the mitoTEMPO antioxidant to patient-derived iPSC-CMs efficiently regulated mitochondrial ROS production and sarcomere organization to rescue the pathological phenotype of Barth syndrome (Wang et al., 2+ handling, leading to CM hypertrophy and dysfunction. Therefore, restoring mitochondrial function by shutting down the exceeding mechanotransduction pathway is a potential therapeutic strategy for cardiovascular diseases. In this review, we highlighted the biological processes of mechanotransduction pathways in regulating mitochondrial function in response to mechanical stress. We also provided a brief description of gene therapy delivery modes used to deliver treatment based on mechanical stress–induced mitochondrial dysfunction, providing new integrative insights into cardiovascular diseases. According to current evidence provided by this review on mechanotransduction in mitochondrial hemostasis, the future cut edge should be located in the following several perspectives. The first thing is to get better understanding of how biophysical mechanical stresses regulate mitochondrial function, as present studies revealed several molecular signals downstream of integrin-FAK–myosin–laminA/C pathway involved in nuclear mitochondrial genes expression. However, mitochondria are found to bind with F-actin, so that the changes of mitochondria are unclear when faced with the physical stress directly. Second, the biochemical sensor–related signaling is really limited to describe, especially the crosstalk between physical and chemical changes under exceeding mechanical stress. At last, to attenuate the abnormal mechanical stress to restore normal mitochondrial function aiming to reverse CMs' phenotype is a promising way. However, the risks of high dosage of AAV administration need to be noticed, and modified strategy is urgently required to be induced to reduce the supplement of AAV and achieve the same therapeutic goal at the same time. In generally, there is still a long way to obtain detailed understanding of mechanotransduction pathway in regulating mitochondrial function and to make targeting treatments.In summary, biophysical and biochemical sensing pathways establish the fundamental mechanotransduction pathways that impact the regulation of mitochondrial homeostasis. Shear stress, cyclic stretch, and static strain are three major mechanical forces CMs are subjected to. Exceeding mechanical stress impairs mitochondrial function through regulating MQC, ATP production, ROS accumulation, and CaHL and PY were primarily responsible for the Biological function of mechanical stress in cardiomyocytes, Molecules involved in mechanotransduction in CMs, and The common understanding of mitochondrial homeostasis sections, respectively. YQ and YY were primarily responsible for the Mechanical cues contribute to the maintenance of mitochondrial homeostasis, and Targeting mechanotransduction pathways to restore mitochondrial function sections, respectively. DZ and YL contributed equally to organize and edit the article. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The Wnt signaling pathway plays important roles in organ development and disease processes. Secreted frizzled-related protein 2 (sFRP2), a vital molecule of Wnt signaling, can regulate cardiac development and cardiovascular disease. Recent studies have suggested that sFRP2 is not only an antagonist of the canonical Wnt signaling pathway, but also has a more complex relationship in myocardial fibrosis, angiogenesis, cardiac hypertrophy and cardiac regeneration. Here, we review the role of sFRP2 and Wnt signaling in cardiac development and cardiovascular disease. The heart is one of the most essential organs for maintaining the normal operation of activities of daily living, and the development of the embryo heart and cardiovascular disease are complex processes. Exploring the regulatory mechanisms that underlie these processes will allow a deeper understanding of cardiac development and provide new therapeutic targets for cardiovascular disease.The Wnt signaling pathway is important for maintaining the homeostasis of embryonic development, and participates in the regulation of biological processes that include cell proliferation, differentiation, apoptosis, and cell localization The proteins of secreted frizzled-related protein (sFRP) family are antagonists of the Wnt signaling pathway, and sFRP2 is considered to be the most potent wg), the segment polarity gene of fruit flies, and the mouse proto-oncogene int-1. In 1982, Nusse et al. first discovered a Wnt gene in the study of a viral transcriptional mechanism in the mouse mammary gland that was found to transmit growth and development signals between cells Wnt genes have been found in mammals. Wnt proteins bind receptors on the cell membrane in an autocrine or paracrine manner to regulate target gene expression. The Wnt signaling pathway is widely involved in early embryo development, cell proliferation, migration, tumorigenesis, metastasis, and stem cell growth regulation 2+/ PKC pathway The term “Wnt” is derived from a combination of two words: wingless (Fz) receptor to form a large complex on the cell surface with low-density lipoprotein receptor-related protein (LRP)5/6 to activate scattered protein (Dsh/Dvl), and affect the distribution of intracellular axis protein Axin. Therefore, inhibiting the phosphorylation activity of GSK3-β leads to stable accumulation of β-catenin in the cytoplasm c-Myc and cyclin D1  have been identified in mammals The sFRP family has long been considered a class of antagonists of the Wnt signaling pathway. Five members of sFRP family  pathway, without affecting the canonical Wnt pathway Fzd5) also plays an important role during angiogenesis promoted by sFRP2. Endothelial cell tubes fail to form when Fzd5 is silenced in endothelial cells sFRP2 exerts pro-angiogenic effects through activation of non-canonical Wnt/CaIt is also generally understood that sFRP2 can exert an angiogenic effect through the above pathways in a wide variety of human tumors, including angiosarcoma, prostate cancer, renal cell carcinoma, lung cancer, and pancreatic cancer Cardiac hypertrophy is an adaptive response of heart under many pathological states such as pressure overload and β-adrenergic stimuli The HyperGEN study used linkage analysis in siblings with hypertension to determine that genetic variation in sFRP2 was associated with left ventricular hypertrophy The limited regenerative potential of cardiomyocytes has made cardiac regeneration therapy by stem cell transplantation a hotspot in cardiovascular disease research Sfrp2 can abolish the aforementioned benefits Bcl-2, inhibit cardiomyocyte apoptosis, and improve cardiac function Mesenchymal stem cells (MSCs) are believed to be a promising candidate stem cell for cardiac therapy in vivoApart from MSCs, sFRP2 also has multiple effects on the properties of other types of stem cells. Aubert et al. found that sFRP2 can promote the differentiation of embryonic stem cells into neural progenitors by antagonizing the canonical Wnt signaling pathway 2+/NFAT pathway, while without affecting the canonical Wnt pathway. Furthermore, sFRP2 has been shown to downregulate the canonical Wnt signaling pathway, and protect cardiomyocytes from pathological hypertrophy. In cardiac regeneration therapy, sFRP2 can improve the survival of stem cells in the adverse environment, and enhance the therapeutic effects for myocardial infarction. Therefore, we believe that sFRP2 is a promising therapeutic target for multiple pathophysiological conditions of cardiovascular disease. However, the specific mechanisms of sFRP2 in these pathophysiological processes require further exploration. Interventions in the Wnt signaling pathway using modified sFRP2 expression may be a promising area of research for heart diseases.SFRP2 plays an important role in the process of heart development and a various cardiovascular pathophysiological condition. In myocardial fibrosis, sFRP2 may play a bidirectional effect depending on the different stage of disease and concentration. Subsequent studies had shown that sFRP2 can promote angiogenesis by activating the non-canonical Wnt/Ca


Menstruation occurs in few species and involves a cyclic process of proliferation, breakdown and regeneration under the control of ovarian hormones. Knowledge of normal endometrial physiology, as it pertains to the regulation of menstruation, is essential to understand disorders of menstruation. Accumulating evidence indicates that autophagy in the endometrium, under the regulation of ovarian hormones, can result in the infiltration of immune cells, which plays an indispensable role in the endometrium shedding, tissue repair and prevention of infections during menstruation. In addition, abnormal autophagy levels, together with resulting dysregulated immune system function, are associated with the pathogenesis and progression of endometriosis. Considering its potential value of autophagy as a target for the treatment of menstrual-related and endometrium-related disorders, we review the activity and function of autophagy during menstrual cycles. The role of the estrogen/progesterone-autophagy-immunity axis in endometriosis are also discussed. Menstruation refers to cyclical shedding and regeneration of the endometrium occurring in very few species. It is regarded as an inflammatory event regulated by ovarian hormones Macroautophagy/autophagy is an intracellular self-digestion pathway for the maintenance of homeostasis. This can be found in all eukaryotic cells at basal level and can be further activated by a variety of stimuli, including deprivation of nutrients, infection, hormones, oxidative stress and endoplasmic reticulum stress It is well accepted that autophagy plays a critical role in cancers, embryo implantation and reproduction According to the conventional viewpoint, by exposure to sequential ovarian estrogen and progesterone will cause the endometrium to undergo well-defined cycles of proliferation, differentiation, and shedding (menstruation) on a monthly basis Among the inflammatory mediators stimulated, IL-8 , monocyte chemoattractant protein-1  and cyclooxygenase-2 , are well-studied Hypoxia during menstruation can directly upregulate vascular endothelial growth factor (VEGF) expression in endometrial stromal cells (ESCs) by modulating the NF-κB pathway Autophagy involves five steps, namely, nucleation, elongation, maturation, fusion and degradation by lysosomes. Double-membraned structures first engulf portions of the cytoplasm to form autophagosomes. Autophagosome is initiated by the unc-51-like kinase 1 (ULK1) complex . This complex can be suppressed via mammalian target of rapamycin complex (mTOR). During autophagosomal vesicle elongation and maturation, autophagy-related (Atg) protein 7 and Atg10 promote Atg12 and Atg5 conjugation. Such that, Atg 12 and Atg5 could interact with Atg16L1 to form a dimeric complex (Atg12-Atg5-Atg16L1), which is required for formation of the covalent bond between LC3 and phosphatidylethanolamine (PE). Then LC3 precursor (pro-LC3) converts into LC3-Ι in the action of Atg4, which is activated by Atg7 and transferred to Atg3. The Atg12-5-16L1 complex facilitates the transfer of LC3-Ι from Atg3 to PE to generate microtubule-associated protein 1 light chain 3 (LC3‑II) LC3-Ι, a marker of autophagy, is increased during autophagy induction, both in endometrial glandular epithelial cells (EPCs) and ESCs throughout the menstrual cycle. LC3-I expression is very low in the proliferative phase, but it surges to peak level at the late secretory phase Figure 1) et al. showed that the expression levels of LC3-II and cleaved caspase-3 increased significantly in normal ESCs cultured with estrogen and progesterone when compared with those cultured with estrogen alone  In vitro experiment also reported the autophagy level in endometrial cells treated with estrogen alone (in vitro imitation of the proliferative phase) was low and was upregulated after the supplementation with progesterone (in vitro imitation of early-to-mid secretory phase) or following the removal of estrogen and progesterone  During the normal human secretory phase ESCs, estrogen is a potent suppressor of autophagy while progesterone can significantly stimulate the activities of autophagy in vitro and in vivoIn addition to ESCs, a large number of EPCs also reside in the endometrium, especially during the secretory phase of the menstrual cycle. The regulation of autophagy in normal EPCs by ovarian steroids has not been reported so far, which may be due to the fact that the primary normal EPCs are very difficult to obtain and culture. Nevertheless, some insights can still be obtained from studies on autophagy in uterine corpus endometrial carcinoma (UCEC) cells. Consistent with estrogen's inhibitory effect on the autophagy of ESCs, our previous studies also show that estrogen significantly restricts autophagy of UCEC cells Figure 1).Therefore, it may be hypothesized that, with estrogen and progesterone withdrawal, there should be higher levels of autophagy in ESC and EPC due to the absence of powerful estrogen-mediated autophagy inhibition , namely ERα, ERβ and G-protein coupled estrogen receptor (GPER) According to a number of recent studies, progesterone and progestins can be particularly important as inhibitors of mTOR in various cell types 2. Notably, overloaded reactive oxygen species (ROS) are generated due to the lack of oxygen, nutrient starvation, mitochondrial toxins and oxidative stress, resulting in organelle damage and the production of toxic substances et al. have reported that estrogen and progesterone withdrawal stimulated PGF2α production through ROS-induced NF-κB activation in human ESCs Figure 2).Hypoxia-inducible factor-1 (HIF-1), a transcription factor which regulates cellular response to hypoxia, is expressed exclusively during the secretory and menstrual phases, suggesting hypoxia would occur in the endometrium late in the menstrual cycle Figure 2). However, the specific mechanism has yet to be clarified.Several studies have identified the AMPK/TSC/mTOR signaling pathway as the classical signaling pathway regulating autophagy in vitroIn response to progesterone, endometrial stromal fibroblasts acquire epithelioid characteristics, such as a highly developed endoplasmic reticulum required for greater secretory function, substantial lysosomes, abundant glycogen and lipids droplets in their cytoplasm Figure 1) Under the cyclic regulation of estrogen and progesterone, leukocytes populations within the endometrium vary considerably across the menstrual cycle, and the mass infiltration of immune cells is closely associated with embryo implantation and menstruation +T cell migration and differentiation 2 and up-regulation of chemokines may be involved in these processes .It has been reported that adipose mTORC1 suppresses PGs signaling and beige adipogenesis via the CREB-regulated transcription coactivator 2 (CRTC2)-COX-2 pathway Macrophages are present throughout the menstrual cycle but display a substantial increase in number during the perimenstrual phase in response to progesterone withdrawal brightCD16-), which is usually found near the endometrial glands and spiral arteries, increase in number from the proliferative phase and peak in the late-secretory phase Typically, NK cell numbers increase dramatically from post-ovulation (around LH+3) to few days prior to menstruation During the early proliferative phase of the menstrual cycle, neutrophils and eosinophils are scarce in the human endometrium. However, their numbers significantly increase when reaching into premenstrual period The spontaneous periodic apoptosis during the menstrual cycle is essential for maintaining the normal structure and function of the endometrium, while cell autophagy plays a critical role in the apoptosis of human endometrial cells at different phases of the menstrual cycle Figure 3). Among these, macrophages and uNK cells secrete MMP-7 and MMP-9, resulting in the apoptosis of smooth muscle cells and endothelial cells + macrophages are abundant in the human endometrium during tissue breakdown and repair. As scavenger cells, macrophages can eliminate apoptotic cells in the uterus and endometrial cells in the peritoneal cavity following the retrograde flow of menstrual blood Under the regulation of endometrial autophagy, a large number of infiltrating leukocytes accelerate endometrial exfoliation and become the main force for removing cellular debris, apoptotic and necrotic cells  +/CD163+) promote tissue remodeling and anti-inflammatory reactions due to reduced proinflammatory cytokine secretion and increased endocytic clearance capacity et al. demonstrated that the hypoxia-autophagy axis induced VEGFA expression in peritoneal mesothelial cells A high level of autophagy during late menstruation facilitates M2 macrophages polarization to participate in efficient repair and remodeling  highCD16- NK cell differentiation Yet, the reduced autophagy and abnormal immune responses in endometriosis are not conducive to the removal of endometrial debris due to impaired immune functions. As previously mentioned, NK cells are involved in the removal of menstrual debris and endometrial fragments that are likely to reach the peritoneal cavity by retrograde menses. Differing from uNK cells in normal endometrium increased remarkedly from the proliferative phase with highest levels in the late secretory phase, uNK cells in ectopic lesions remained significantly low throughout the cycle 2 in PF 2 on promoting cell autophagy 2/autophagy signaling. These M2 macrophages can facilitate the remodeling of ECM and neovascularization of lesions Notably, the elevated levels of peritoneal IL-8 due to defect autophagy enhance the FasL-induced apoptosis in activated T lymphocytes, contributing to the immune-privileged environment around the endometriotic implants and supporting their survival Programmed cell death 4 (PDCD4), a well-documented tumor suppressor, could inhibit tumor cell proliferation, migration and invasion at the transcriptional and translational levels As discussed previously, the estrogen/ progesterone-autophagy-immunity axis is involved in the shedding of the endometrium, tissue regeneration and the defense mechanism during menstruation. Aberrant levels of autophagy are responsible for menstrual-related and endometrium-related diseases in vivo interventions in humans is not plausible. Thus, further well-designed studies would be required to fully understand the mechanisms underlying the pleiotropic roles of this axis in order to enrich our knowledge of physiological mechanisms associated with menstruation and pathological events such as menstrual dysfunction, endometriosis, and miscarriage.Given that autophagy is a dynamic process, static measurements could lead to misinterpretation of the result. Additionally, studies on the mechanism of menstruation are limited, due to the lack of suitable animal models (menstruation does not take place in rodents) and


MSN2, which is the transcription factor gene in response to stress, is well-known to increase the tolerance of the yeast Saccharomyces cerevisiae cells to a wide variety of environmental stresses. Recent studies have found that the Msn2 is a feasible potential mediator of proline homeostasis in yeast. This result is based on the finding that overexpression of the MSN2 gene exacerbates the cytotoxicity of yeast to various amino acid analogs whose uptake is increased by the active amino acid permeases localized on the plasma membrane as a result of a dysfunctional deubiquitination process. Increased understanding of the cellular responses induced by the Msn2-mediated proline incorporation will provide better comprehension of how cells respond to and counteract to different kinds of stimuli and will also contribute to the breeding of industrial yeast strains with increased productivity.Overexpression of  Moreover, environmental fluctuations may cause protein-related damages, such as the inhibition of enzyme activities, destabilization of cellular structures, and instability of chemical gradients, which eventually result in cell disruption. Thus, protein quality control and protein homeostasis are essential prerequisites for stress responses. Under harsh stresses, cells also undergo the systematic downregulation of energy-producing and energy-consuming processes in order to enter into a quiescent state, often accompanied by a dynamic shift in the central metabolic pathways that convert nutrients into energy and biomass. Cells possess tight and precise regulation systems to coordinate all the changes that are interconnected at those different levels.Cells of microorganisms are frequently exposed to a variety of stresses in their environment, such as prolonged nutrient starvation, free radicals and toxic molecules, imbalances in osmotic pressure and pH level, and non-optimal growth temperatures   are reduced, yeast cells reprogram the modes of energy metabolism from fermentation to respiration, a process termed a diauxic shift, in order to maximize the efficiency of energy production  controlled by the yeast-specific transcription factors Msn2 and Msn4 (Msn2/4) includes responses to various stresses, such as oxidative stress, osmotic shock, glucose starvation, high ethanol concentrations, temperature upshift, and freezing stress  and reside in the cytoplasm. Once yeast cells are challenged by environmental perturbations, Msn2/4 are rapidly dephosphorylated and translocated into the nucleus , SOD1 and SOD2 (for superoxide dismutases), and PRX1 and TSA2 (for thiol peroxidases) , which is converted to proline by the P5C reductase Pro3  superfamily consists of 24 permease proteins whose function is to transport amino acids and other amines into the cells , a toxic analog for proline, is used in both fundamental and applied researches, as it has been proven to be beneficial to study the cellular metabolism and the production of macromolecules in both prokaryotes and eukaryotes , and l-canavanine (Can), which are known to be the analogs of proline, phenylalanine, and arginine, respectively  cells could be due to the increased incorporation of AZC, since higher AZC levels were detected in MSN2-OE cells than that observed in wild-type cells  in MSN2-OE cells with the strain CAY29 background conferred higher resistance to AZC  or arginine  and is classified into ubiquitin-specific-protease (USP) family caused depletion of free cellular ubiquitin (Chernova et al., UBP6 and other DUB genes was not significantly changed in MSN2-OE cells, indicating that Msn2 does not repress UBP6 and other DUB genes transcriptionally (Mat Nanyan et al., UBP6 in yeast cells makes them more susceptible to the toxicity of translational inhibitors such as cycloheximide and the toxic arginine analog, Can, which are caused by a deficiency of free ubiquitin (Chernova et al., UBP6, UBP3 as well as OTU1 genes aggravates the growth inhibitory effect of toxic amino acid analogs in yeast cells (Mat Nanyan et al., UBP6 led to the most remarkable sensitivity toward AZC, OFP, and Can (Mat Nanyan et al., MSN2-OE cells, suggesting that the excess level of Msn2 and Ubp6 deficiency confer a common phenotype of defective resistance against toxic amino acid analogs. The combination of the MSN2 overexpression and UBP6 disruption resulted in growth similar to that seen with Δubp6 cells in the presence of low concentrations of AZC (Mat Nanyan et al., MSN2 and UBP6 increased the resistance to AZC, OFP, and Can (Mat Nanyan et al., ubp6 cells and were mostly localized on the plasma membrane, similar to what was observed in MSN2-OE cells (Mat Nanyan et al., S. cerevisiae in order to withstand various fluctuating growth conditions.Ubiquitination and deubiquitination are involved in the control of endocytic degradation of amino acid permeases localized on the plasma membrane (Springael and André,  of Gnp1 . AlthougMSN2 has been shown to increase the tolerance of yeast cells for various kinds of stress conditions, such as oxidative and freezing stresses. In addition, we shed light on a potentially important link, namely that the inhibition of endocytic degradation mediated by Msn2 plays an essential role in regulating the proline homeostasis under stress conditions, evidently caused by the loss of function of DUBs. Further investigations will elucidate the profound relationship among Msn2, DUBs, and Gnp1 in the regulation of proline homeostasis, which may serve as a foundation to engineer more robust industrial yeast strains, which might be advantageous in fermentation industries.In this mini-review, we discuss the current understanding of the stress-responsive transcription factor Msn2 and the stress response in yeast. In particular, the overexpression of NM and HT substantially and intellectually contribute to this work, and reviewed and approved the final version of manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


PIK3CA gene.  Megalencephaly-capillary malformation (MCAP) is a rare overgrowth syndrome caused by postzygotic activating mutations in the  To illustrate the benefits of gingival biopsy in the genetic diagnosis of overgrowth syndromes. PIK3CA sequencing was performed on DNA extracted from gingival biopsies, blood, and saliva.  Gingival biopsy was performed on a 13-year-old patient and a 16-year-old patient with MCAP and who suffered from periodontal disease. PIK3CA mutations were found in the gingival biopsies with an allelic frequency of 22% and 35%, respectively, while they were undetectable in blood or saliva. The genetic diagnosis of MCAP through detection of PIK3CA somatic mosaicism in a periodontal biopsy is unprecedented.  Pathogenic p.Glu365Lys and p.Glu545Asp PIK3CA mutation, the composite embryologic origin of periodontium and its high fibroblast cell content make it an ideal target for molecular analysis in overgrowth syndromes, and multidisciplinary approach including paediatric dentists should be encouraged. In addition, our clinical findings suggest that periodontal disease is part of the MCAP phenotypic spectrum and should be systematically investigated. Considering the tissue distribution and level of somatic mosaicism for  PI3KCA-related overgrowth spectrum (PROS) includes MCAP syndrome and several other disease entities with clinical overlap . MCAP syndrome (MIM#602501) is characterized by megalencephaly, capillary malformations, asymmetric growth, polymicrogyria, polydactyly, and syndactyly [Megalencephaly-capillary malformation (MCAP) syndrome is a rare congenital condition belonging to the segmental overgrowth disorder group caused by somatic mutations in the RTK/PI3K/AKT/mTOR growth pathway . In thisPIK3CA gene (locus 3q26.32) encodes the 110 kD (p110) alpha subunit of the phosphatidylinositol 4,5-bisphosphate 3-kinase (PI3K) involved in intracellular signalling. The PI3K signalling pathway is important for many cellular activities including cell growth, division, migration, angiogenesis, and survival. These functions make PI3K an essential protein in the development of tissues, including brain tissue and blood vessels. Some PIK3CA mutations have oncogenic potential (MIM∗171834). In MCAP syndrome, PIK3CA mutations lead to an increased p110 alpha subunit activity that results in constitutive PI3K signalling [The Clinically, MCAP syndrome may result in a disproportionate head due to an enlarged brain and capillary malformations  on the skin of the midline face, trunk, and limbs. The oral manifestations of this syndrome are not yet described; the following two case reports provide a description of the particular aspect of periodontal disease in children with MCAP.To illustrate the interest of gingival biopsy in the diagnosis of such syndrome, we report here the cases of a 13-year-old patient who consulted in Toulouse University Hospital in March 2015 for bleeding gums. Her medical history revealed a clinical diagnosis of MCAP not confirmed by genetics.Her medical drug therapy consisted of analgesics for chronic pain in the lower right limb with lymphatic malformations. Lymphatic drainage and the wearing of a compression garment at night were implemented. She also had a slight inequality in the length of her lower limbs (2-3 mm), for which orthopaedic soles were made.During early childhood, the lymphatic malformations of the right hemiface resulted in bites on the internal surface of the overgrown cheek. A lipoaspiration was thus carried out in 2006 followed by wearing of a contention mask for 1 month in order to decrease the volume of the right cheek. The Institutional Review Board approved this study, and the treatment was performed after obtaining written consent from the parents.Exobuccal examination revealed slight right side dominant facial asymmetry. We did not observe cutaneous capillary malformations on her face. The endobuccal clinical examination  revealedExplanations to the patient and her parents about the illness Oral hygiene motivation and educationAntibiotic treatment (amoxicillin and metronidazole)Removal of orthodontic applianceTooth scalingClassical periodontal therapy was established, including the following:After reevaluation, root planing was performed in two sessions under conscious sedation with nitrous oxide (due to the age and anxiety of the patient).A gingivectomy was also performed, and samples of periodontal tissue in the most affected area  and saliva were sent for genetic analysis .After 6 months, the clinical improvement was notable but gingival inflammation persisted, with induced bleeding and a budding and erythematous appearance of certain papillae, especially in the mesial papilla of the right upper lateral incisor .A literature search was conducted for intraoral periodontal and mucosa abnormalities associated with MCAP syndrome. No articles reporting intraoral lesions in MCAP subjects were found. However, capillary malformations of the face were reported , 4.PIK3CA gene by a next-generation sequencing process with confirmation of the variants by the Sanger method [Gingival biopsy was performed on the mesial papilla of the right upper lateral incisor which was the most budding and erythematous area. Genetic analysis was carried out on the r method .PIK3CA gene analysis was performed on DNA from gingival biopsy, blood, and saliva using next-generation sequencing (NGS) with confirmation by the Sanger method. The pathogenic mutation NM_006218.2:c1093G>A (p.Glu365Lys) was found only in the gingival biopsy with an allelic ratio of 22%  gingival appearance and an increased frequency of gingival and periodontal pathologies.Additional case reports will be required to determine the set of associated periodontal and mucosal pathologies found in MCAP patients.PIK3CA gene in a periodontal biopsy is unprecedented. However, it is not the first time that a mosaic mutation of the PIK3CA gene is found in an intraoral sample. McDermott et al. identified a mosaic mutation of the PIK3CA gene (c2176G>A) in the DNA of a permanent tooth. Among the DNA of the blood, hair, and dental samples, it was the dental DNA that showed the highest degree of mosaicism [AKT1 gene, which occurs in the same intracellular signalling pathway as the PIK2CA gene, was identified in an odontogenic cyst of a patient with Proteus syndrome [Concerning the first case, the identification of mosaic missense mutations in the  gene c216G>A in tsyndrome .PIK3CA mutation, the complex embryologic origin of periodontium and its high fibroblast cell content make it an interesting target for molecular analysis in overgrowth syndromes. For the same reason, the periodontium is more interesting to detect genetic variations than buccal mucosa biopsy or saliva collection. Moreover, greater attention should be paid to hypertrophic gingival lesions in such syndromes.Gingival biopsy is rarely used for the diagnosis of genetic diseases unlike blood or skin samples . NeverthIn those two case reports, the dental surgeon helped in the diagnosis of MCAP. As part of the multidisciplinary care of these patients, a deeper involvement of paediatric dentists should be considered for the treatment of any associated periodontal abnormalities as well as for taking samples for genetic analysis and diagnosis.