Transcription of the ribosomal RNA genes (rDNA) that encode the three largest ribosomal RNAs (rRNA), is mediated by RNA Polymerase I (Pol I) and is a key regulatory step for ribosomal biogenesis. Although it has been reported over a century ago that the number and size of nucleoli, the site of ribosome biogenesis, are increased in cancer cells, the significance of this observation for cancer etiology was not understood. The realization that the increase in rRNA expression has an active role in cancer progression, not only through increased protein synthesis and thus proliferative capacity but also through control of cellular check points and chromatin structure, has opened up new therapeutic avenues for the treatment of cancer through direct targeting of Pol I transcription. In this review, we discuss the rational of targeting Pol I transcription for the treatment of cancer; review the current cancer therapeutics that target Pol I transcription and discuss the development of novel Pol I-specific inhibitors, their therapeutic potential, challenges and future prospects. In “western countries” cancer is now responsible for the majority of disease related deaths each year . A signiThe transformation of normal cells into cancer cells requires the gradual acquisition of certain characteristics, coined “the hallmarks of cancer” ,7. TheseRiBi takes place in sub-nuclear domains termed nucleoli that have long been linked to cancer with the enlargement and increase in the number of nucleoli per cell being used for over a century as a marker of malignancy . More coThe 80S ribosomes are composed of two subunits: small subunit (40S) that binds and scans mRNA  and the cis-regulatory elements and the coding region of the 47S precursor RNA transcript, that is processed to give rise to 18S, 5.8S and 18S rRNAs, and a long intergenic spacer  or “nucleolar surveillance pathway” (NSP) to trigger a variety of downstream effects including cell cycle arrest and apoptosis. The best studied NSP mechanism leads to activation of the p53 (TP53) pathway. During times of steady state rRNA transcription MDM2, an E3 ubiquitin ligase, ubiquitinates p53, priming it for proteasome degradation. When a key stage of RiBi, rRNA synthesis, is acutely disrupted through any one of a variety of mechanisms , “free” Ribosomal Proteins (RP) RPL5 and RPL11 (as part of the 5S-RNP particle) which are no longer being incorporated into the ribosome, bind and sequester MDM2 thus preventing it from ubiquitinating p53. This in turn results in the rapid accumulation of p53 protein which transcriptionally regulates target genes involved in cell cycle arrest, apoptosis and genome stability . AdditioRiBi upregulation, modulated by oncogenes like MYC, is essential for the survival and proliferation of cancer cells . The obsHere we review the current knowledge of the effect of the non-selective and selective inhibitors on rRNA transcription . By analA large variety of antineoplastic drugs are currently being used as therapeutics for cancer. These drugs can be classified accordingly to their mode of action into four major groups: Alkylating agents, that covalently attach alkyl groups to other molecules including DNA; Antimetabolites, that mimic the structures of normal metabolites; Antibiotics that bind/intercalate with DNA preventing RNA synthesis; and Plant alkaloids that prevent cell division . InvariaAlkylating agents: Platinum-based alkylating compounds, like Cisplatin and its analogues Carboplatin and Oxaliplatin, are widely used chemotherapeutics in a variety of cancers, particularly those with high mortality rates like small cell lung carcinoma, ovarian and head and neck squamous cell carcinoma [arcinoma . Platinuarcinoma . Althougarcinoma ). Recentarcinoma ,48. Cisparcinoma ,50, resuAntimetabolites: Antimetabolites are drugs that mimic the structure of cellular metabolites and inhibit specific enzymes causing DNA synthesis inhibition [hibition . Severalhibition .5-FU and its metabolites can be incorporated into RNA and DNA as well as bind to the active site of the Thymidine Synthetase enzyme, inhibiting its activity and consequently affecting the production of dTTP, leading to a deficit in Thymidine required for DNA synthesis. The combination of these attributes leads to both RNA and DNA damage and consequently the death of cancer or normal cells . Thus, Methotrexate can bind to the enzyme Dihydrofolate Reductase and prevents it from reducing folate. If folate is unavailable, the Thymidine Synthetase enzyme is incapable of converting Uracil into Thymidine leading to a deficit in Thymidine required for DNA synthesis. Burger et al. showed that treatment of cells with Methotrexate reduced Pol I transcription and leads to disruption of the nucleolus suggesting activation of NSP .Antibiotics: Intercalating agents are antibiotics that exert their chemotherapeutic function by insertion between DNA bases. The downstream effects of the drugs in this category varies from induction of DNA damage  and Topoisomerase poisoning . All these drugs have long been used as chemotherapeutic agent for the treatment of multiple types of cancer [f cancer .Actinomycin (ActD), also known as Dactinomycin, intercalates with both double and selective single stranded DNA, with a preference for GC rich sequences . IntercSimilarly to ActD, Mitomycin C (MMC), when in its reduced form, preferentially intercalates with GC-rich regions and alkylates DNA causing DNA damage  and ultiDoxorubicin and Mitoxantone also intercalate with DNA but they act as Topoisomerase II poisons. Topoisomerase II enzymes remove DNA supercoils by a mechanism involving a transient break on both strands of DNA to allow strand passage. Intercalation with DNA leads to the formation of DNA adducts that prevent the Topoisomerase from re-ligating the DNA strands and therefore causes double strand breaks that cannot be repaired . The DNAPlant Alkaloids: Plant alkaloids are natural products that interfere with cell cycle and DNA synthesis [ynthesis . One exaynthesis .Camptothecin, Topotecan and Irinotecan inhibit Type I Topoisomerases, that removes DNA supercoils by making a break in a single strand of DNA . TopoisoEllipticines are a family of planar plant alkaloids that can penetrate the cell by diffusion and inhibit Topoisomerase IIα. Several derivatives of the parent compound ) have been approved for use in clinical trials, but they have not progressed after stage one or two due to adverse side effects of the treatment . HoweverThe knowledge that Pol I transcription was frequently upregulated in cancer combined with the observation that many existing chemotherapeutic drugs possess substantive anti-Pol I transcription activity led to the realisation that drugs which selectively target Pol I transcription might constitute a new class of anticancer therapies with increased potency and reduced toxicity than non-selective inhibitors of RiBi. In early 2008, Cylene Pharmaceutical commenced high throughput screens for small molecules that could disrupt Nucleolin/rDNA G4 complexes and identified a series of fluoroquinolone derivatives, the lead compound of which was termed CX-3543 . SubsequCX-3543: CX-3543, also known as Quarfloxin  have moderate levels of Pol I transcription that steadily increase through progressively lineage-restricted progenitors, while mature haematopoietic cells are characterized by low levels of rDNA transcription ,84,85. TCX-5461 showed therapeutical efficiency in mice bearing MYC-driven lymphomas , models Interestingly, in more recent studies an additional mode of action for CX-5461  was propCX-5461 has recently completed a Phase I clinical trial in patients with advanced haematological cancers . In thisCX5461 is also currently in Phase 1/2 clinical trial for breast cancer (NCT02719977). Similarly to the clinical trial in haematological cancers, CX-5461 was well tolerated and efficiency was observed in patients with homologous recombination-deficient tumours .BMH-21: BMH-21 was the most potent of six compounds identified through a high content screen as being able to induce p53 activation. BMH-21 is a DNA intercalator however it does not activate the cellular DNA damage response [response . It mediresponse . A recenresponse  proposedTwo contradictory reports evaluated the potential of BMH-21 to intercalate and stabilize G4 DNA structures. Musso et al.  showed bThe collective pre-clinical data on specific Pol I transcription inhibitors developed so far clearly demonstrate their therapeutic potential for the treatment of cancer. However, despite the clear benefits of these new drugs, CX-5461 is the only inhibitor of its class to have entered clinical trials . While pioneering and extremely promising, CX-5461 is associated with additional activities, like DNA damage , which possibly contributes to its efficacy, toxicity profile and resistance mechanisms.Therefore, the search for improved inhibitors or the second generation continues. A new chemical series of Pol I transcription inhibitors is currently under research by Pimera Inc. . The lead molecule, PMR-116, shows improved chemical properties and higher efficiency compared to CX-5461 and is currently being prepared for first-in-human clinical trials with cancer patients. Further research is also being performed on other hits identified in the screen for BMH-21 .Although Pol I transcription inhibitor drug development is gathering pace, significant questions remain about their clinical application including patient stratification; what predicts resistance to Pol I transcription therapy; and what drugs might combine synergistically with Pol I transcription inhibitors to increase patient survival and overcome resistance. Finally, thought needs to be given to other steps in RiBi that might be amenable to drugging and whether the downstream signalling, and thus therapeutic effects, would be the same or differ to those Pol I transcription inhibitors currently developed.The biggest challenges of cancer therapies that target housekeeping processes, like RiBi, is the identification of the types of cancer and groups patients that would best respond to the treatment. The use of the appropriate cohort of patients is crucial for the success of clinical trials. Testing on subjects not likely to respond will undermine the value of Pol I transcription inhibitors and potentially lead to reduced efficacy. Identification and validation of biomarkers is therefore a priority for the development of all new therapeutics, including Pol I transcription inhibitors.Numerous publications have considered stratification of patients based on RiBi levels or nucleolus size  with mixA potential biomarker for stratification of patients for Pol I transcription inhibition therapy is the presence of a functional p53 protein. Although NSP can also occur via a p53-independent pathway, the most common, and perhaps stronger, response to cellular stress is achieved by the p53- dependent NSP. Cellular stress leads to altered RiBi/Pol I transcription allowing for the stabilization of p53 and the activation of its downstream pathways leading to cell death or a block in cell proliferation ,42,43. AThe expression level of the oncoprotein MYC (cMYC and N-MYC) is another potential biomarker to predict therapeutic response to Pol I transcription inhibitors. MYC expression is deregulated in a large proportion of cancers and its overexpression is linked with poor prognosis ,100. ThrCancer cells without a functional homologous recombination mechanism, are known to be more sensitive to therapies that produce DNA double strand brakes, like radiotherapy and Topoisomerase II inhibitors . ConsequThe ATRX  gene is mutated in approximately 5% of all cancers with the highest incidence in glioblastoma, neuroendocrine pancreatic cancer (53% with mutations in DAX/ATRX), non-small lung carcinoma and sarcoma ,113,114.KDM4A (JMJD2A) is a member of the KDMA histone demethylase family and, among other targets , it also removes the repressive chromatin mark H3K9me3 . TherefoPre-existing resistance and/or acquired resistance to therapy by circumvention of the drug targeted pathway is a common occurrence during cancer treatment. This is achieved by multiple mechanisms including pre-existing or acquired mutations in the direct drug target or downstream pathways or upregulation of alternative compensatory pathways. The use of a combination of drugs allows targeting of multiple pathways or one pathway at multiple levels, thereby reducing the chances of resistance development, is a common approach used to attain more durable responses in patients. Rational approaches to combinatorial therapies have been tested for Pol I transcription inhibitors with promising results ,122.The ability of MYC to drive protein synthesis is essential for cancer proliferation and can be inhibited at different levels from ribosome production to protein translation. Targeting protein synthesis at different levels is therefore predicted to improve therapeutic efficiency. Indeed, treatment of MYC-driven B-cell lymphoma with CX-5461, via inhibition of rDNA transcription, in combination with Phosphoinositide 3-kinase (PI3K), AKT and mammalian target of rapamycin complex (mTORC) inhibitors, disrupting the recruitment of ribosomes to the mRNA, lead to increased tumour cell death and prolonged survival . HoweverInhibition of ATM/ATR pathways in murine models of AML  or poly These results emphasize the fact that drug combinations need to be tailored to specific types of cancers and/or mutations. Therefore, several specific drug combinations are being assessed.It is clear that selectively inhibiting Pol I transcription is an effective way to treat cancer without inducing general DNA damage and therefore would reduce toxicity and the probability of developing resistance. Pol I transcription can be inhibited at different stages and, because of the need of a multi-protein interaction to initiate and maintain Pol I transcription, a multitude of potential therapeutic targets can be envisioned. Fine mapping of the human Pol I complex and its associated proteins  using crystallography or cryo-electron microscopy should allow us to design new inhibitors in a more targeted way, for example by fragment-based design. Using these techniques, we should be able to identify the protein interaction surfaces that would be more amenable to disruption by the presence of a small molecule. Together with the knowledge of the specific role of each of these proteins on Pol I transcription, i.e., knowing which proteins to target in order to obtain higher specificity and lower toxicity will be an invaluable asset. RRN3 provides one such example. RRN3, is necessary for the interaction between the SL-1 complex and PIC, mediated by the Pol I subunit RPA43. Studies demonstrate that disruption of the interaction between these complexes by CX-5461 leads to selective inhibition of Pol I transcription  suggestiPerhaps one of the most attractive Pol I components for therapeutically targeting is UBF. UBF is a cytoarchitectural transcription factor that is essential for rDNA transcription. UBF binds at the rDNA promoter, across the Pol I transcribed portion of the rDNA repeat and remodels the chromatin at the rDNA locus to maintain permissive chromatin allowing for the fine-tuning of Pol I transcription levels in response to environmental changes ,124,125.For many years, RiBi was considered exclusively a housekeeping process, and not a potential target for anti-cancer therapy. However, a paradigm shift has been observed over the past decade. It has become evident that selective inhibition of Pol I transcription has the potential to become a novel and efficient therapeutic approach in the fight against cancer. Moreover, it was convincingly demonstrated  that man


Orofacial clefts (OFCs) are the most common congenital birth defects in humans and immediately recognized at birth. The etiology remains complex and poorly understood and seems to result from multiple genetic and environmental factors along with gene–environment interactions. It can be classified into syndromic (30%) and nonsyndromic (70%) clefts. Nonsyndromic OFCs include clefts without any additional physical or cognitive deficits. Recently, various genetic approaches, such as genome-wide association studies (GWAS), candidate gene association studies, and linkage analysis, have identified multiple genes involved in the etiology of OFCs.This article provides an insight into the multiple genes involved in the etiology of OFCs. Identification of specific genetic causes of clefts helps in a better understanding of the molecular pathogenesis of OFC. In the near future, it helps to provide a more accurate diagnosis, genetic counseling, personalized medicine for better clinical care, and prevention of OFCs. Orofacial clefts (OFC) are the most common congenital malformations in the orofacial region causing a significant personal and social encumbrance.The prevalence of OFCs ranges from 1 in 700 to 1,000 newborns worldwideThe prevalence of OFC varies according to geographical location, ethnicity, race, gender and socioeconomic status,The frequency of occurrence of OFC differs with regard to gender and side of clefting. Cleft lip is more common in males at a 2:1 male to female ratio, whereas a cleft palate is more common in females.The development of lip and palate begins during the fourth week of gestation where migrating neural crest cells combine with mesodermal cells to establish facial primordia which consist of five different processes  derived from the first pharyngeal arch.The secondary palate begins to develop in the seventh week of embryogenesis; the maxillary processes initially outgrow as palatal shelves which move toward each other vertically. After proper growth they settle at a horizontal position above the tongue which entails much extracellular remodeling.In summary, a variety of cellular mechanisms such as cell proliferation, cell migration, cell growth, cell fusion, apoptosis, EMT, and extracellular remodeling are involved in a coordinated manner during the development of lip and palate. Therefore, disruption in the gene/s involved in these processes during lip and palate development may lead to an OFC.Although the familiarity of OFC has long been noted, Fogh-Andersen was the first to provide the evidence for genetic factors contributing to the etiology of CLP from family-based studies where it was observed that the siblings of CLP patients had an increased frequency of cleft lip with or without cleft palate.The etiology of OFCs involves genetic factors, environmental influences, and gene–environment interactions, all contributing to its susceptibility. Scientific literature evidence suggests that environmental factors such as maternal tobacco smoking and alcohol consumption, antiepileptic medications, maternal folate deficiency, infections, consanguinity, and geographical location are risk factors for NS cleft lip and palate (NSCLP).Advances in genetics and molecular biology techniques have discovered multiple genes and loci associated with CLP. This article provides an overview of the genes implicated in the etiology of NSCLP. Identification of specific genetic variation contributing to NSCLP has led to an increase in our understanding of the molecular pathogenesis of OFC.SATB2gene mutation. Recently, using salivary miRNAs showed that theSATB2genes are involved in the development of the cleft palate and lip development.Special AT-rich sequence-binding protein (SATB2) is a DNA binding protein which binds with nuclear matrix attachment regions. It is involved in transcription regulation and chromatin remodeling process. In an animal study, mouse SATB2 is strongly expressed in the developing cleft palate and is similar to the human SATB2 protein.BCL3gene has also shown a strong association with NS orofacial clefts (NSOCs).BCL3gene with NSCLP was found in multigenerational families of Indian population.B-cell leukemia/lymphoma 3 (BCL3) is a proto-oncogene, acts as a transcriptional co-activator through NF-kappa-B target genes and located on 19q13.2.DLXgene family containing a homeobox transcription factor which plays an important role in craniofacial development and palatogenesis. It is located on chromosome 17q21.33 and causes orofacial cleft 15 (OFC15) and cleft lip/palate. In an animal study, theDLXgenes caused cleft palate showing the importance of these genes in craniofacial morphogenesis.Distal-Less Homeobox 4 (DLX4) belongs toPAX9) is a member of the paired box (PAX) family of transcription factors and contains a paired box domain, an octapeptide, and a paired-type homeodomain. It plays a critical role during neural crest and fetal development. PAX9 is located on chromosome 14q13.3, consists of five exons, and associated with the formation of the teeth and palate.Paired Box Gene 9  used unmixed NS CPO and NS CLO subtypes suggested that PAX9 is a strong genetic factor for NS CPO in the Chinese population. Mutation analysis of PAX9 SNPs rs12885612 and rs12881248 revealed thatThe netrin 1 (OMIM: 601614) located on chromosome 17p13.1 is a family of laminin-related secreted proteins. Its functions include axon guidance and cell migration in the central nervous system, angiogenesis, and semicircular canal formation.NTN1 is expressed at high levels in cells that will come together to form a fusion plate, a prerequisite for the formation of semicircular canals. In netrin 1 mutant mice, fusion plate formation is severely affected, and it stimulates proliferation of the periotic mesenchymal cells which then push the epithelial cell walls together to form the fusion plate.A GWAS included 1,409 case-parent trios by several research groups with samples of Asian or European ancestry from Europe, the United States, China, and the Philippines found NTN1 role in the etiology of NSCLP.T-box 22(TBX22) gene encodes transcription factors involved in the regulation of developmental processes and plays a major role in human palatogenesis. It contains eight coding exons. TBX22 cause X-linked cleft palate and is characterized by isolated cleft palate and ankyloglossia.TheGenome-wide linkage analysis showed the association of TBX22 role in the development of NSCLP.Poliovirus receptor-like 1 (PVRL1), also known as NECTIN1 (nectin cell adhesion molecule 1) belongs to the nectin subfamily of immunoglobulin-like adhesion molecules that involve cell–cell adhesion. It plays a vital role in the organization of adherens junctions and tight junctions in epithelial and endothelial cells.PVRL1gene resulting CLPED in families from Israel and Brazil.PVRL1gene were identified in Turkish NSCLP patients.In animal experiments, PVRL1 expressed at the medial edge epithelium of the palatal shelves and the skin surface epithelium locations that corresponded to the clinical phenotypes of CLPED and in humans, mutations of theCLPTM1genes suggested that a regulatory element in this gene region get affected and which is responsible for the development of orofacial clefts.Cleft lip and palate-associated transmembrane protein 1 (CLPTM1) is a multipass transmembrane protein that regulates GABA-A receptors  and modulates inhibitory synaptic strength. It is located on chromosome 19q13.3 and plays a role in T-cell development. Mutation of(MAFB)gene encodes a basic leucine zipper (bZIP) transcription factor that plays an important role in the regulation of lineage-specific hematopoiesis. It is located on chromosome 20q11.2, consists of a single exon and spans approximately 3 kb.MAFBgene reported causing multicentric carpotarsal osteolysis syndrome and Duane retraction syndrome 3 with or without deafness.The MAF bZIP Transcription Factor BMAFBgene SNP (rs13041247) is associated with NSCL/P risk in different population; however, this association is not significant in East Asian or Caucasian populations,Several GWAS in European and Asian populations identified the role of MAFB in NS CLP.A variety of genetic approaches have identified several genes located on different chromosomes, contributing to etiology of NSCLP. Advances in genetics and molecular biology techniques have led the way to the discovery of genetic variation involved in NSCLP. Genes such as FGFR1, TFAP2A, GSTT1, receptor-like tyrosine kinase (RYK), and ABCA4 have also been found to be associated with NSCLP.FGFR1gene in NSCLP families and suggested that the FGF signaling pathway may contribute to as much as 3 to 5% of NS cleft lip or palate.FGFR1gene in Chinese Kallmann syndrome males with cleft lip/palate.Riley et al assessed the genes involved in the fibroblast growth factor (FGF) signaling pathway and identified the functional impairment in theRYKgene were also found to be associated with orofacial clefting.ABCA4gene association with NSCLPFurther, mutations in theGenetic studies provide insight into the multiple genes involved in the etiology of OFCs. Identification of specific genetic causes of clefts helps for better understanding of the molecular pathogenesis of OFC. In the near future, it helps to provide a more accurate diagnosis, genetic counseling, personalized medicine for better clinical care and prevention of OFCs.


To the Editor—Many US hospitals have recently adopted policies mandating universal masking of all staff, visitors and patients. Universal masking is particularly important in preventing transmission to and from individuals who are asymptomatic or presymptomatic for coronavirus disease 2019 (COVID-19).1 The Centers for Disease Prevention and Control (CDC) estimates that ~35% of severe acute respiratory coronavirus virus 2 (SARS-CoV-2) cases are asymptomatic.2 In addition, healthcare professionals (HCPs), patients, and visitors with atypical or very mild symptoms may more readily transmit SARS-CoV-2 in healthcare facilities without masking policies.3 Risk of exposure to SARS-CoV-2 in nonclinical areas within healthcare facilities may be overlooked.4 We recently surveyed 50 community hospitals within the Duke Infection Control Outreach Network and found that 90% of these hospitals had also adopted universal masking policies. However, we also determined that actual compliance with universal masking policies was suboptimal, particularly among staff outside of clinical care settings, including administrative offices, shared work rooms, and break rooms. Poor compliance in these shared spaces led to known exposures in some of these hospitals, leading to employee furloughs, a substantial burden of contact tracing, and unnecessary anxiety for exposed individuals. Here, we discuss our perception and understanding of the etiology of poor compliance with universal masking policies in healthcare settings, and we discuss proposed solutions as well.The incidence of COVID-19 among HCPs decreased significantly after our health system adopted a universal masking policy. Unmasked exposure to another HCP rather than exposure to known infected patients resulted in most of the COVID-19 cases among staff after implementation of this policy.5 HCPs commonly perceive their risk of contracting COVID-19 from an infected patient to be higher than the risk of exposure to an asymptomatic coworker. Ironically, HCPs spend more time in close proximity to their coworkers than infected patients. A recent study demonstrated that <5% of exposed HCPs tested positive for SARS-CoV-2 despite exposure to an infected patient without adequate personal protective equipment (PPE), although most HCP attribute greater risk to this type of exposure.6 The propensity among HCPs to perform inaccurate risk assessments has been seen with other basic infection prevention measures such as hand hygiene.7Overall, HCP compliance with protective measures such as universal masking often correlates with the level of risk they perceive. Individuals are more likely to comply with recommended prevention measures if they perceive themselves to be at higher risk of harm in a particular situation or setting.5Earlier this year, public health authorities pointed out a lack of evidence related to the use of universal masking by the general public to prevent acquisition of SARS-CoV-2. Later, a member of the World Health Organization (WHO) stated in June 2020 that asymptomatic spread of SARS-CoV-2 is ‘very rare.’ The WHO quickly modified and clarified this statement by stating that asymptomatic spread is incompletely understood even though it actually occurs, contributing to ongoing confusion. Furthermore, a few high-ranking political leaders and millions of citizens routinely ignore the current recommendation to use face coverings in indoor settings and when in close proximity with others. Inconsistent, contradictory and unclear advice from public health authorities has contributed to widespread confusion about the utility of universal masking in preventing the spread of SARS-COV-2 (response efficacy).9 In our opinion, this guidance is confusing and adds an unnecessary emphasis on the use of additional PPE by HCPs when in direct contact with patients and does not place emphasis on the need for universal masking of patients when staff are in close proximity to patients.The CDC recently updated their exposure guidelines and issued a new “frequently asked question” on May 29, 2019, recommending the use of eye protection when caring for patients in areas of “moderate to substantial community transmission [of SARS-Cov-2],” even if COVID-19 is not suspected.10 This “fatigue” among HCPs may be potentially related to their long work hours, required interactions with other team members throughout the day, the burden of wearing additional eye protection and uncomfortable or poor-quality masks.COVID-19 fatigue, a term that describes drift in following preventative measures as this pandemic goes on, is an important cause of poor compliance with policies related to universal masking.11 We need to work closely with HCPs to better understand the root causes for poor masking compliance and to identify and remove barriers to doing the right thing. Simple solutions such as educational campaigns on the rationale for masking, creation of a mask committee comprised of key stakeholders from various worker types to serve as champions, making physical changes to the environment to facilitate distancing, offering better quality masks, as well as suitable and accessable alternate locations that allow for physical separation to occur while HCPs are unmasked during breaks, will likely lead to improved compliance.For effective behavioral change, wearing a mask must become a habit for HCPs in all shared spaces inside and outside the workplace, outside of their immediate household and when appropriate physical distancing is not possible.12Finally, we need clear, simple, and consistent messaging from public health authorities for successful implementation of universal masking policies. Our goal should be to focus on the simple message of universal masking to prevent the transmission of SARS-CoV-2. Healthcare epidemiologists and public health professionals need to learn the art of salesmanship during these times because the message itself, though important, is only as good as the leader that presents it to the public.


This study aims to describe the technical success of the micropuncture technique, which is performed in placement of tunneled hemodialysis catheters in patients with central venous occlusion and limited access.A total of 25 patients with central venous occlusion and in need of catheter placement for hemodialysis between 2012 and 2018 were included in this study and analyzed retrospectively. Technical success was defined as the placement of tunneled dialysis catheters with optimal position and function.Internal jugular vein access in 16 patients (14 right and 2 left) and right subclavian vein access in 3 patients were successfully performed in placement of the tunneled dialysis catheter. Although internal jugular and subclavian vein access was attempted bilaterally, the procedure failed in 6 patients. The overall technical success of recanalization of the occluded central veins was 76% (19/25). No minor or major complications were encountered.Tunneled dialysis catheter placement through the occluded internal jugular and subclavian veins with the micropuncture technique is effective and safe in patients with limited vascular access. The recanalization of the occluded conventional access routes should always be kept in mind to allow for the preservation of vascular accesses for future requirements. The use of tunneled hemodialysis catheters, especially in patients with end-stage renal disease and with limited access for surgical fistulas, is increasing owing to its practical placement and management [1]. Central venous catheterization is one of the most common procedures performed in interventional radiology units because of its low complication rates, high technical success, and long-term patency . A history of prior multiple catheter placements and long-term indwelling catheters can result in central venous stenosis or occlusion, and this can result in limited vascular access for dialysis sessions [3]. Internal jugular veins, subclavian veins, and femoral veins are accepted as the conventional access sites for tunneled catheters [2]. In case of occlusion or adversity while accessing the veins mentioned above, alternative or unconventional venous access routes, including translumbar inferior vena cava, transhepatic inferior vena cava, small venous collaterals, and recanalization of occluded central veins, can be used [4–8]. Also, the sharp puncture technique for recanalization of chronic central venous occlusions has been described for patients that could not be accessed with a guidewire [9]. Recanalization of the conventional veins in patients with central venous occlusion and limited access are primarily preferred owing to their superficial location and because it is easily accessible, whereas unconventional access routes such as translumbar inferior vena cava and hepatic veins are accepted as technically challenging and time consuming [2].The purpose of this study is to describe the technical success of the micropuncture technique, performed in placement of tunneled hemodialysis catheters in patients with central venous occlusion and limited access.A total of 25 patients with central venous occlusion and in need for catheter placement for hemodialysis between 2012 and 2018 were included in this study. Patients from hemodialysis centers were mainly referred to us to manage dysfunction of the current tunneled dialysis catheters or to place a new dialysis catheter in order to use it again. All patients had a history of end-stage kidney disease, multiple previous catheter placements, and a necessity for urgent dialysis. Low-molecular weight heparin was started in all patients before the procedure. The recanalization procedure was performed immediately due to the urgency of the need for dialysis. Doppler ultrasonography (US) was primarily used to detect the patency of the veins before planning the recanalization and catheter placement . The age of the thrombus was determined with respect to the findings in Doppler US, whereas fluoroscopy during the procedure or contrast-enhanced computed tomography before the procedure were used in detection of thrombus length. Anechoic or minimal echogenic thrombus with venous distension was defined as acute, whereas echogenic thrombus with normal or minimal reduced calibration of vessel was defined as subacute thrombosis. Sonographic findings in the chronic stage of thrombosis are decreased vein diameter, diffuse thickening of the vein wall, and echogenic fibrous synechiae or bands [10]. Occlusions extending to the middle part of the brachiocephalic vein were classified as short and those extending to the vena cava superior were classified as long segment.The main inclusion criteria for recanalization of occluded central veins were having no surgical upper limb dialysis options  and limited access sites  for catheter placement. Patients with chronic central vein occlusions accompanying collateral vessels, uncorrected coagulation disorders, and severe orthopedic disorders preventing supine position were excluded from the study.All patient data included in our study, such as demographic information and clinical findings, were obtained from medical records and from our procedure form. Informed consent forms were signed before treatment, and the ethics committee approved our study design.Patients with coagulation parameters within normal limits  were accepted for the procedure. All procedures were performed in the interventional radiology unit under the guidance of ultrasonography  and fluoroscopy . Patients were put in supine position on the C-arm fluoroscopy table, and sterile conditions were provided. Neither sedation nor general anesthesia was used during the procedures; only local anesthetic agent (10-mL lidocaine 2%) was applied to the puncture region. Just above the clavicles, a 21-gauge (G) needle with an echogenic tip was inserted into the thrombosed segment of the internal jugular vein under US guidance, enabling the most parallel route to the longitudinal axis of the vessel as possible. In patients with failure of access via internal jugular veins bilaterally, a subclavian vein puncture was performed. A 0.018-inch × 45-cm nitinol guidewire was delivered through the needle under fluoroscopy, advanced into the patent central vein (vena cava superior) and right atrium after crossing the thrombosed segment. Alternatively, a 0.018-inch hydrophilic guidewire  was used in cases when there was difficulty in crossing long-segment occlusions. A 4-French (F) sheath  was advanced into the thrombosed internal jugular vein through the microguidewire. After withdrawing the guidewire, the distal tip of the sheath was confirmed as being in the patent lumen by injection of contrast medium. In order to provide extra strength and stability, a 0.035-inch J–tip guidewire  was delivered through the 4-F sheath and preferably advanced to the inferior vena cava in order to avoid serious cardiac arrhytmia. Gradual dilation was performed with the use of 8-F, 10-F, 12-F, and 14-F dilators before placement of the peel-away sheath of the dialysis catheter. In patients exhibiting difficulty in gradual dilation, a balloon with 5-mm diameter was used for the luminal patency needed for catheter placement. After maintaining vascular access with a peel-away sheath, the procedure was concluded in the same way as a routine tunneled dialysis catheter placement. At the end of procedure, the function and position of the catheters  were checked under fluoroscopy, and patients were discharged with recommendations for post-procedure catheter care. The recanalization and catheterization steps are demonstrated in Figures 2A–2F. No routine antibiotic prophylaxis was administered during the procedures.Technical success was defined as the placement of the tunneled dialysis catheter in optimal position and function. We define the optimal position as being the location where the tip of the catheter placed within the junction of the vena cava superior and right atrium. Clinical success was defined as resumption of normal dialysis for at least 1 session. The CIRSE classification system was used in defining the complications [11]. Numeric data were presented as average, standard deviation, median, maximum, and minimum, while categorical data were presented as number and percentage. Statistical data editing and analysis were performed using SPSS .A total of 25 patients  with central vein occlusion and a history of multiple dialysis catheter placements were retrospectively analyzed in this study. The age of the thrombus in the occluded segment demonstrated by Doppler US were acute in 3 patients (12%) and subacute in 22 patients (88%). The length of the thrombus in the occluded segment demonstrated by fluoroscopy during the procedure or contrast enhanced computed tomography before the procedure were short segment in 17 patients (68%) and long segment in 8 patients (32%). In our study, external jugular vein accesses were not feasible for cannulation due to insufficient vessel diameter or thrombosis. Considering femoral vein access: 8 patients (32%) had a nonfunctioning femoral catheter due to chronic femoral vein or vena cava inferior occlusion, 4 patients (16%) had an infection of femoral catheters, and 13 patients (52%) refused femoral catheterization due to difficulty of care and high risk of infection. In all patients, right internal jugular vein puncture was performed as the first choice due to its straight course to the right atrium. Internal jugular vein access in 16 patients (14 right and 2 left) and right subclavian vein access in 3 patients were successfully performed in placement of the tunneled dialysis catheter. Although internal jugular and subclavian vein accesses were attempted bilaterally, recanalization failed in 6 patients (24%). In 4 of the 6 patients, central venous catheters were placed via femoral vein despite high infection risk and 2 of the 6 patients were referred for surgical lower limb dialysis access. Overall, technical success in recanalization of the occluded central veins was 76% (19/25). Considering the age of occlusion, successful recanalization rates in acute and subacute thrombus were calculated as 100% (3/3) and 72.7% (16/22), respectively. The procedure was successful in 50% (4/8) of patients with long-segment occlusion and 88.2% (15/17) of patients with short-segment occlusion. During the procedures, no minor or major complications were encountered. Patients were followed up with respect to catheter dysfunctions and survival. Median follow-up time in 19 patients with successful recanalization and catheterization was 7.95 months . During follow-up, catheter dysfunction and catheter-related infection were observed in 5 (26.3%) and 4 patients (21%), respectively. In these cases, catheters were exchanged through a guidewire from the same access point, and patients were discharged with functional central venous catheters. The primary patency rate was 78.9% at 6 months. Four patients died due to comorbid diseases during follow up. Patients’ demographic, central venous occlusion, and catheterization-related data are shown in the Table.Several methods, such as native arteriovenous fistula, arteriovenous grafts, and central venous catheters, have been defined for hemodialysis process [12]. Central venous tunneled catheter placement in patients with end-stage renal disease and whose surgery offered no opportunity to create an upper limb arteriovenous fistul/arteriovenous graft or presented the required time for fistula maturation is essential for continuous dialysis sessions [2]. Central venous catheterization is of critical importance not only for patients with end-stage renal disease but also for all chronic patients needing easy venous access for medication or nutrition [13]. Central venous stenosis, thrombosis, and infection are accepted as inevitable results of prolonged hemodialysis, especially in patients with comorbidities . The role of interventional radiology has been expanding in parallel with the increasing need for dialysis worldwide, either in the placement of dialysis catheters and management of the catheter dysfunctions or for catheter-related infections. In the literature, several venous access routes have been defined in which to place a tunneled dialysis catheter. Although there is no consensus on which vascular access is best, clinical history, venous anatomy, and preference of clinician and operator are the decision-making parameters during planning [2]. In our interventional radiology unit, internal jugular vein access is preferred as the first choice to reduce several complications such as thrombosis and dysfunction. Beside the conventional venous access routes, salvage techniques, such as translumbar inferior vena cava access, transhepatic inferior vena cava access, catheterization of small venous collaterals, and recanalization of occluded veins, were reported [4]. As each access option has several advantages and disadvantages, operator experience is the main factor in determining the optimal salvage technique.Recanalization of the occluded veins in catheter placement was first described in 1996 by Ferral et al. in 6 patients. In their study, a femoral vein approach was primarily used to get venous access. Before recanalization of the occluded innominate and subclavian veins, a 10-mm Amplatz Goose Neck snare was placed at the subclavian and supraclavicular area position as a target. Under fluoroscopy guidance, a 21-G needle was directed to the snare. Following the confirmed position of the needle, a 0.018-inch guidewire was advanced into the snare and through-and-through access was provided. With the support of through-and-through access, the central venous catheter was placed after dilation of the occluded vein [16]. In our study, no femoral venous access was required during the procedure in patients with successful recanalization of occluded internal jugular or subclavian veins. The micropuncture technique used in upper central veins is feasible even in patients with occluded femoral veins and vena cava inferior. In 2016, Too et al. reported about a technique called REBORN, which entailed the recanalization and balloon-oriented puncture for reinsertion of a dialysis catheter in nonpatent central veins. In their technique, a balloon is used as a target of the puncture to access the central vein. [17]. In their study, an angioplasty balloon is used as part of their technique; however, we needed angioplasty only in patients having difficulty in gradual dilation. The micropuncture technique for vascular access offers a small puncture site, less vascular injury, and the opportunity to initiate multiple punctures with the help of a 21- (G) needle when compared with the standard 18-G vascular access needle [18]. Besides the diameter of the micropuncture needle, the echogenic tip design also improves visualization under ultrasonography guidance, increasing the success rate and minimizing complications [19].Other reasons for our 21-G needle preference are that the 0.018-inch guidewire in the micropuncture set allows it to be easily manipulated due to the short length of the guidewire, and it has better torque capability while crossing thrombosis.Studies concerning the comparison of the micropuncture needle (21-G) and regular needles (18-G or 19-G) have been frequently performed during femoral arterial access . Gutzeit et al. reported slightly a lower number of pseudoaneurysms in the micropuncture group; however, no significant decrease in complication rates was observed [20]. During central venous cannulation, inadvertent carotid artery puncture with standard needles might lead to a mortal hemorrhage in the neck region. Micropuncture needles reduce hemorrhagic complications, even in patients with uncorrected coagulopathy [18].Retrograde recanalization of central venous occlusions via femoral vein route offers a lower risk of vascular complications, owing to the anatomic features of the femoral puncture site, as well offering a straighter path when compared with antegrade recanalization via internal jugular or subclavian veins [22]. Complications related with the canulation of upper central veins include: hemothorax, pneumothorax, carotid artery injury, mediastinal hematoma, and cardiac tamponade. As in our study, antegrade recanalization of upper central veins with the help of a micropuncture needle prevents such complications; thus, it can be used safely in patients with end-stage renal disease and limited vascular access for dialysis.The major limitations of our study are retrospective design, the small number of patients, and lack of long-term patency rates and overall survival rates. The small number of patients included in the study is due to the fact that patients with central venous occlusion and limited vascular access are less commonly seen than patients undergoing standard central venous catheterization. In conclusion, tunneled dialysis catheter placement through the occluded internal jugular and subclavian veins with the micropuncture technique is effective and safe in patients with limited vascular access. We believe that maximum care and time should be spent for patients with end-stage renal disease due to limited vascular access sites for catheter placement. The recanalization of the occluded conventional access routes should always be kept in mind to allow the preservation of vascular access for future requirements.This research received no specific grant from any funding agency in the public, commercial, or not-for-profit sectors.


Both RT and IN have been successfully targeted with potent antiretroviral drugs to inhibit these enzymes and treat patients infected with HIV-1, the causative agent of acquired immunodeficiency syndrome (AIDS). Despite decades of research, the molecular mechanisms of reverse transcription and integration in vivo are not fully characterized, in part because cell-free assays that efficiently and faithfully recapitulate the processes in the context of a virion have not been available.1 One of the key differences between this study and previous efforts to develop efficient ERT assays was the addition of IP6, a highly negatively charged compound that was recently shown to bind to the central pore of capsid hexamers and stabilize the HIV-1 capsids.2 Careful quantitative PCR analysis of reverse transcription products indicated that in ∼50% of the viral cores reverse transcription progressed to a late stage called plus-strand strong-stop DNA transfer, confirming that the conditions of the ERT assay supported efficient reverse transcription  assay, Christensen and colleagues used purified virions, gently lysed them with a pore-forming melittin peptide from honey bee venom, and empirically optimized buffer, salt, deoxynucleotide triphosphates (dNTPs), rNTPs, inositol hexokinase (IP6), and cell extracts.Next, the authors examined how capsid stability affects reverse transcription by determining how capsid mutants that increase capsid stability, moderately decrease stability, or severely decrease stability affect reverse transcription efficiency. In addition, they determined the effect of IP6 concentrations on the efficiency of reverse transcription for wild-type, as well as the mutant capsids. The results showed that IP6 concentrations close to the estimated cellular concentrations yielded peak reverse transcription products for wild-type virions, suggesting that there is optimal capsid stability for supporting reverse transcription. The optimal IP6 concentrations were lower for hyperstable capsids and higher for less stable capsids, providing additional evidence that optimal capsid stability is important for efficient reverse transcription. Furthermore, treatment of capsids with capsid-binding inhibitor GS-CA1 disrupted capsids and inhibited reverse transcription, indicating that maintaining intact capsids is critical for efficient DNA synthesis.3Electron cryotomography (ECT) and subtomogram lattice mapping were employed to visualize individual cores at the subunit level. Under the conditions of the ERT assay, the morphologies of the individual capsids varied greatly at a time when most of the late reverse transcription products had been synthesized (8–10 h). While most cores (56%) remained largely intact, a significant proportion of cores (36%) had morphologies spanning a continuum from partially disassembled to almost completely disassembled capsid shells. The partially disassembled capsids were often missing local areas of CA hexamers, indicating that patches of CA hexamers were lost without complete disassembly of the capsid. In many of the partially disassembled capsids, the authors often observed viral DNA strands exiting and looping back into the capsid. The DNA loops extruding out of the partially disassembled capsids were estimated to be quite short, ranging from 0.5 to 1.5 kb, suggesting that most of the 9.7-kb DNA remained tightly packed inside the remaining capsid shell. In the absence of dNTPs, a majority (82%) of the capsids remained largely intact, indicating that reverse transcription promotes capsid disassembly. It remains unclear whether the capsid disassembly initiates only after completion of viral DNA synthesis, whether the partially disassembled capsids represent abortive reverse transcription, or whether reverse transcription can be completed after the capsid is partially disassembled. Nevertheless, one can envision that the authors have captured the uncoating process, with each partially disassembled capsid potentially representing a different stage of uncoating on the path toward complete capsid disassembly. Consistent with a previously proposed model, the authors suggest that the formation of dsDNA, which is relatively rigid compared to RNA, increases internal capsid pressure, resulting in local disruption of the capsid.4 Strikingly, integration required the addition of cell lysate, indicating that one or more host factor(s) is needed to facilitate the reaction.At least some of the reverse-transcribed dsDNA synthesized in the ERT assay could integrate into a plasmid DNA target using the virion-incorporated IN. The integration was dependent on the completion of viral DNA synthesis and sensitivity to IN inhibitors. The authors also documented concerted integration, and nearly all of the events involved the expected 5′ target site duplication. Integration also required capsid disassembly as previously reported for HIV-1 capsids in the nuclei of infected cells.4 The results of these robust cell-free assays, and those from cell-based assays by us and others have gratifyingly converged to show that reverse transcription primarily occurs in the context of an intact viral core.5 It is anticipated that the cell-free ERT assays described by Christensen et al. will yield many insights into the early stage of HIV-1 replication for years to come.1These studies describe efficient in vitro reconstitution of HIV-1 reverse transcription and integration. The results show that reverse transcription evolved to be an efficient process, that an intact capsid is essential for viral genome replication, and that capsid disassembly is a requirement for integration. Historically, it has not been possible to address these questions in infected cells because most viral cores that enter cells do not lead to productive infection and it has been difficult to quantify the amount of CA associated with the infectious viral cores. Our recent studies show that the viral cores remain intact or nearly intact in the nuclei of infected cells and undergo a rapid uncoating event (<20 min) near the site of integration shortly before integration.


Granule cells (GCs) are the most numerous cell type in the cerebellum and indeed, in the brain: at least 99% of all cerebellar neurons are granule cells. In this review article, we first consider the formation of the upper rhombic lip, from which all granule cell precursors arise, and the way by which the upper rhombic lip generates the external granular layer, a secondary germinal epithelium that serves to amplify the upper rhombic lip precursors. Next, we review the mechanisms by which postmitotic granule cells are generated in the external granular layer and migrate radially to settle in the granular layer. In addition, we review the evidence that far from being a homogeneous population, granule cells come in multiple phenotypes with distinct topographical distributions and consider ways in which the heterogeneity of granule cells might arise during development. In this review article, we address how granule cells (GCs) develop: their origins in the upper rhombic lip  and the elaborate migrations and amplifications that lead GCs to become the most populous neurons in the brain, and secondly the embryological origins of granular layer (GL) heterogeneity. Given the space constraints we do not review synaptogenesis and circuit formation; recent reviews may be found in Leto et al.  and LackThe adult cerebellar cortex has four distinct layers. Superficially lies the molecular layer, a cell-poor structure rich in GC axons, Purkinje cell (PC) dendrites, and their synapses. Next is the narrow PC layer comprising a monolayer of PC somata. Beneath the PC layer is the GL, densely packed with GC somata. Each of these three cellular layers also has distinct populations of inhibitory interneurons. The three cellular layers overlie the white matter tracts, which comprise the cortical afferent and efferent axon pathways  to form physically separate and molecularly distinct clusters by ~E18 : fortunately, either way, it abbreviates as EGL. The EGL was first described thoroughly by Hess , together with excitatory cerebellar nuclei neuron progenitors and glutamatergic UBC progenitors, are generated dorsally in the upper rhombic lip , where t by Hess  and then by Hess , and was by Hess  to gener by Hess . As they by Hess .via GC axons to the PCs, which are the sole efferent projection of the cerebellar cortex  input from incoming mossy fiber afferents , and lonr cortex . While tr cortex ! It is nr cortex . Therefor cortex .in situ hybridization  rostrally and rhombomere 2 (Hoxa2+) caudally. In the arealization of the URL, Gbx2 plays an important role through the antagonism of Otx2 expression  whose mutated gene was found to be Lmx1a  and this pathway is likely involved in the production of the crucial progenitor gene Atoh1  to the adult population in the GL (~P25), there is a ~3,000× amplification! As  the EGL . Scant il12 gene , 6F,G exl12 gene , 6H,I, el12 gene . Fetal cdeleted in colorectal carcinoma  and subsequent EGL maintenance (E18–P20) both require contacts with the basement membrane, involving both the basal endfeet of radial glial progenitors and fibroblasts of the pia mater. In 1985 Hausmann and Sievers , and thereby promote GCP proliferation  protein family converts SHH signals into gene expression regulation . ATOH1 activates ciliogenesis by transcriptionally regulating Neurod1  show a medial fusion defect at E16 but no other obvious histological abnormalities  is reduced both in Zic1−/− and Zic1+/–; Zic2+/kd cerebella, and the expression of mitotic inhibitor p27 (Cdkn1b), which inhibits GCP proliferation  driven by the Cntn2 promoter delays but does not arrest GCP proliferation in the EGL  are expressed in the inner part of the EGL, where GCPs exit the cell cycle and start to differentiate. Vitronectin promotes phosphorylation of cyclic-AMP responsive element-binding protein (CREB), whose overexpression is sufficient to induce GC differentiation even in the presence of SHH  and other factors involved in neuronal migration and connectivity  tag GCs that occupy more caudal territories  do not move into the anterior cerebellum in mutants in which the early-born (Lmx1a-) GCs are absent, again pointing to the presence of an AZ/CZ restriction boundary (see below). Likewise, in the embryonic primordium, the Ebf2 gene, encoding a helix-loop-helix transcription factor involved in cerebellar cortical development  has pronounced GL defects focused on the posterior vermis , critically regulates GCP adhesion during EGL exit via the junctional adhesion molecule JAM-C  glial cells and their Bergmann fibers  expressed on migrating GCs  mice  E3 ubiquitin-protein ligase complex  and ROSA26 controls [Sey/Sey⟷Gt(ROSA)26Sor], the AZ/CZ boundary is apparent, together with the second boundary in lobules VII/VIII  mutant the anterior cerebellum is agranular with a transition to normal at the AZ/CZ boundary: the phenotype seems intrinsic to GCPs : Ackerman et al., disabled . This was first described for NADPH/nNOS expression  populate anterior segments of the EGL  into four distinct molecular subgroups—WNT-dependent, SHH-dependent, Group 3, and Group 4 (reviewed in Northcott et al., Pax6 mutant small eye (Sey, Engelkamp et al., Finally, the array of parallel fibers is highly conserved through vertebrate evolution and thus is important, but its development remains poorly understood. The mechanisms that restricts the extension of parallel fibers on the frontal plane (e.g., Berglund et al., In conclusion, we believe that GC development remains a vital research field, not only because it involves the origin of over half of all neurons in the brain but also for its far-reaching implications in developmental biology, human genetics, oncology, and regenerative medicine.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.


This review focuses on the importance of developing meat products with healthier lipid content and strategies such as the use of structured lipids to develop these enriched products. The review also conducts a critical analysis of the use of vibrational spectroscopy as a tool to further these developments. Meat and meat products are extensively recognized and consumed in the world. They are an important nutritional contribution in our diet. However, their consumption has also been associated with some negative consequences for health due to some of its components. There are new trends in the design of healthy meat products focusing mainly on improving their composition. From among the different strategies, improving lipid content is the one that has received the most attention. A novel development is the formation of lipid materials based on structured lipids such emulsion gels (EGs) or oil-bulking agents (OBAs) that offer attractive applications in the reformulation of health-enhanced meat products. A deeper interpretation is required of the complicated relationship between the structure of their components and their properties in order to obtain structured lipids and healthier meat products with improved lipid content and acceptable characteristics. To this end, vibrational spectroscopy techniques (Raman and infrared spectroscopy) have been demonstrated to be suitable in the elucidation of the structural characteristics of lipid materials based on structured lipids (EGs or OBAs) and the corresponding reformulated health-enhanced meat products into which these fat replacers have been incorporated. Future research on these structures and how they correlate to certain technological properties could help in selecting the best lipid material to achieve specific technological properties in healthier meat products with improved lipid content. The global dynamics of the production and consumption of meat and meat products has evolved rapidly as the result of changing lifestyles and nutritional ideologies among part of the population. As a result, it is important to address several different aspects regarding the quality of meat and meat products, particularly nutritional , safety and sustainability aspects.Regarding the nutritional aspects of meat and meat products, different strategies have been explored to optimize the composition of these products to make them healthier and to bring them in line with health recommendations and nutritional guidelines promoted by public bodies in response to new consumer demands.Of the different strategies to improve meat and meat product composition, lipid content optimization has attracted the most attention owing to health recommendations . This tySuccessful development of structured lipid materials with healthier characteristics requires in-depth study of their nutritional, sensorial and technological characteristics, and a broader understanding of the complex relationship between the structure of their components and their properties. A new approach to the understanding of these interrelationships based on an analysis of the conformational modifications taking place within the components of lipid material and reformulated new meat products that affect their properties, merits consideration. In these connections, vibrational spectroscopic techniques such as Raman and infrared spectroscopy are direct and non-invasive techniques which offer an extensive range of possibilities in meat and meat products ,7,13 andBased on the above, this review features a brief description of the novel strategies used to develop meat products with a healthier lipid content based mainly on the use of structured lipids (emulsion gels and oil bulking agents). It also proposes the employment of vibrational spectroscopy (infrared and Raman spectroscopy) to enhance the development of both structured lipids and the meat products which are included as animal fat substitutes. Moreover, the basic concepts of infrared and Raman spectroscopy are reviewed to aid in the understanding of how these spectroscopic techniques are applied.Meat and meat products form part of the diet of many consumers in the world and are a significant source of an extensive range of nutrients essential for healthy development. These foods provide many nutrients such as high-value protein, fatty acids such as conjugated linoleic acid, minerals  and B-complex vitamins. However, meat products also contain processing additives  with negative health implications. This has sparked the development of meat products more in line with health recommendations  using diFrom among the different reformulation processes, improved lipid content  and quantitative terms) is one of the most relevant processes in the development of healthier meat products ,2,3,15 bThe consumption of SFA has been related to the development of cardiovascular disease, obesity, hypertension and some types of cancer . The conDespite this controversy, general consumption recommendations of total and unsaturated fatty acids can be made ,22. BrieTo meet these recommendations and improve the lipid content of meat products, technological strategies generally replace animal fat with different lipids, mainly from plant and marine sources, more in line with health recommendations , higher in monounsaturated (MUFA) and polyunsaturated fatty acids (PUFA), especially from the omega-3 or n-3 family of fats). Efforts are also being made to improve PUFA content, the n-6/n-3 and PUFA/SFA ratios and, where possible, to reduce cholesterol and trans-fatty acids) . This wiDifferent procedures have been used to substitute animal fat with healthier lipids, ranging from the most conventional  to the most novel recently developed techniques such as lipid structuring with EGs or the oil-bulking agents referred to above ,2,3. To Lipid materials created using lipid structuring procedures have obtained a great deal of attention in the context of meat products. These lipid constituents can be employed as fat substitutes in meat products thanks to their solid-like properties simulating animal fat ,2,3. The2+), and enzymes with hydrocolloids [EGs are lipid materials in which emulsions and gels (hydrogels) concur. Formulation of these EGs fundamentally includes a lipid phase , an aqueous phase, an emulsifier  and a gelling process ,2,3. Difcolloids ,23. Partcolloids .Other interesting structured lipids are OBAs based on the dispersion of oil droplets in a continuous aqueous matrix-forming gel . In thesMany studies have been conducted in recent years concentrating on the creation and characterization of EGs and their application in food products , particularly in the elaboration of cooked and fresh meat products  with a healthier lipid content ,15,24,25To better understand the use of vibrational spectroscopy in the development of EGs and OBAs and in meat products with a healthier lipid content, it is necessary to adequately know the basic concepts of these techniques and the interpretation of the spectral results obtained with them.Vibrational spectroscopy, which involves infrared (IR) and Raman spectroscopies, is founded on the transitions concerning quantized vibrational energy states of molecules. In IR spectroscopy, the energy for these transitions is supplied by radiation in the IR regions (mid-IR and near-infrared) of the electromagnetic spectrum . In RamaIR and Raman spectroscopy provide many special benefits in food researches ,27,28,29Infrared and Raman spectroscopy offer relevant data on the structure of the components of meat and meat products  non-invasively and in situ ,30. The Mid-infrared (IR) spectroscopy is the most widely used spectroscopic technique in meat and meat products as it provides fundamental information about protein and lipid structure through well-resolved characteristic infrared bands. In recent years, mid-infrared Fourier transform (FT-IR) spectrometers with attenuated total reflection (ATR) stand out from among the different mid-infrared (IR) spectroscopy techniques employed to analyze the structure of meat and meat products.−1), amide II (1560–1510 cm−1) (−1) which provide information on protein secondary structure . Modifications in the intensity and/or frequency of these infrared bands is indicative of protein structural modifications [−1, whereas bands between 1640 and 1612 cm−1 are usually related with β-sheets.The typical protein bands in the mid-infrared (IR) spectra are amide I (1700–1600 cm10 cm−1)  and amidications ,30. The −1 region and overlapping bands in the 1500–700 cm−1 region. It is worth noting that in the elaboration of meat products with health-enhanced lipid content, infrared studies of the region between 3000–2800 cm−1 are characterized by two strong bands at 2925 and 2854 cm−1 (asCH2) and symmetric (νsCH2) stretching vibrations of the acyl CH2 groups of lipids [The mid-IR spectra of lipids encompass various bands in the 3000–1700 cm854 cm−1  caused rf lipids ,32,33. Mf lipids .Raman spectroscopy offers data on protein structure, mostly by analysis of the amide I and III bands, which are associated with secondary structure. This spectroscopic technique also provides bands due to the environment of some side chains of proteins  and on the local conformations of disulfide bonds and methionine residues, all associated with a tertiary protein structure ,29. Four−1), which involves mostly C=O stretching, and amide III vibrational modes (1225–1350 cm−1)  are amide I (1650–1658 cm50 cm−1) . Amide Im−1) . Nevertheless, one of the most frequently used regions of the Raman spectra in the structural study of lipids in meat and meat products, especially those products with health-enhanced lipid content, is between 2800–3000 cm−1  and IνsCH3/IνasCH3 (I2935/I2890) offer valuable indices to measure lipid packing consequences and establish relative order/disorder of the intermolecular lipid chain [There are also several Raman bands allocated to lipids near 1750, 1660, 1470, 1443, 1306, and 1269 cm000 cm−1  associatid chain ,37.−1) (−1) including the bending (δ) and stretching (ν) vibrations of the O(N)-H..O(N) units, due to interactions of hydrogen bonded to water and protein molecules [Raman spectra also show a broad water band (3100 and 3500 cm−1) , associaolecules ,39.Once this basic knowledge about vibrational spectroscopic techniques has been established, their application on the development of EGs and OBAs and healthier meat products in which these structured lipids are incorporated will be described separately.−1, and between 2600 and 3100 cm−1, were selected for regression models since these contain bands related to lipids. Results showed that Raman spectroscopy is an interesting technique to measure the fatty acid composition of pork adipose tissue with the benefit that this technique is non-invasive and measurements can be completed online [Many researchers have studied the possibilities of vibrational spectroscopy to establish fat content and the fatty acid composition of animal fat  ,41,42,43d online ,43,44,45−1 region indicate that the order/disorder of the oil lipid chain, associated to lipid interactions and droplet size in the emulsion gels, could be important in controlling their textural properties [FT-IR has been employed to study conformational modifications of oils emulsified with proteins  where these proteins are adsorbed in the emulsion formation process ,47. Thisoperties .Polysaccharides, a structured lipid, can be employed either individually or in mixture to form a diversity of gel structures which may be appropriate for immobilizing oil droplets working as oil-bulking agents. Raman spectroscopy has also been employed to determine lipid and polysaccharide interactions in different polysaccharide gels elaborated for use as oil-bulking agents . Raman sThis information is of particular interest given that these lipid materials can help to improve or maintain the quality of the food to which they are added. Particularly important in this regard is their use as animal fat replacers, for instance in the development of health-enhanced meat products without compromising the properties of the final product.−1, particularly the half-bandwidth in the 2922 cm−1 band, has suggested modifications in lipid chain order or lipid–protein interactions. These results underscore the relevance of the stabilizing system employed in the preparation of oil-in-water emulsions as it impacts both the structural and technological characteristics of the reformulated meat products.Vibrational spectroscopic techniques, particularly FT-Raman and FT-IR, provide information about the secondary structure of proteins and the structure of lipids in meat and meat products ,13,27,28−1 region showed that frankfurter reformulated with chia emulsion gels implicate more lipid–protein interactions which correlated significantly with processing loss (mainly loss of water and fat) and the textural behavior of these samples [−1) of the ATR-FTIR spectrum presented greater (p < 0.05) inter- and intramolecular lipid disorder irrespective of the presence of polyphenol in the EG [Lipid materials based on structured lipids such as chia EGs have been applied as animal fat substitutes to develop frankfurters with a health-enhanced lipid profile. In this case, ATR-FTIR spectroscopy was used to study how replacing animal fat with chia EGs affects the structural characteristics of lipids . Results samples . ATR-FTIn the EG .−1 regions and intensity ratios IνsCH2/IνasCH2 (I2850/I2890) and IνsCH3/IνasCH3 (I2935/I2890) revealed that the addition of oil-bulking agents as animal fat replacers in these health-enhanced frankfurters increased lipid acyl chain disorder and implicated more lipid−protein interactions [FT-Raman spectroscopy was also employed to study the structural properties of health-enhanced frankfurters reformulated with structured lipids such as oil-bulking agents to replace fat . Analysiractions .The notion of improving the lipid component in meat products has been gaining attention in recent years giving rise to the development of new reformulation strategies to obtain healthy meat products and optimal technological properties. New procedures to create lipid materials based on structured lipids, such as EGs and OBAs, are especially interesting owing to their solid-like properties and the fact that they confer valuable nutritional and technological properties to meat products into which these materials have been added. EGs and OBAs have been employed successfully as animal fat replacers in the development of different meat products  with healthier lipid contents .Application of vibrational spectroscopy (infrared (IR) and Raman spectroscopy) has significant possibilities and an increasing range of applications in the analysis of structured lipids (mainly EGs and OBAs) and meat products into which these lipid materials have been incorporated for the purpose of achieving healthier lipid contents. The use of vibrational spectroscopic techniques for both lipid materials and health-enhanced meat products have provided us with a greater insight into how the structure of their main components (proteins and lipids) impacts technological properties, mainly texture. This information can be helpful in choosing the most appropriate compounds for lipid materials used as animal fat replacers to obtain specific technological behaviors. Structural information and its correlation with certain technological properties could help in the selection of the most optimal lipid material and other compounds to obtain certain properties in health-enhanced meat products into which they are included.


Skeletal muscle deteriorates with aging, contributing to physical frailty, poor health outcomes, and increased risk of mortality. Denervation is a major driver of changes in aging muscle. This occurs through transient denervation-reinnervation events throughout the aging process that remodel the spatial domain of motor units and alter fiber type. In advanced age, reinnervation wanes, leading to persistent denervation that accelerates muscle atrophy and impaired muscle contractility. Alterations in the muscle fibers and motoneurons are both likely involved in driving denervation through destabilization of the neuromuscular junction. In this respect, mitochondria are implicated in aging and age-related neurodegenerative disorders, and are also likely key to aging muscle changes through their direct effects in muscle fibers and through secondary effects mediated by mitochondrial impairments in motoneurons. Indeed, the large abundance of mitochondria in muscle fibers and motoneurons, that are further concentrated on both sides of the neuromuscular junction, likely renders the neuromuscular junction especially vulnerable to age-related mitochondrial dysfunction. Manifestations of mitochondrial dysfunction with aging include impaired respiratory function, elevated reactive oxygen species production, and increased susceptibility to permeability transition, contributing to reduced ATP generating capacity, oxidative damage, and apoptotic signaling, respectively. Using this framework, in this review we summarize our current knowledge, and relevant gaps, concerning the potential impact of mitochondrial impairment on the aging neuromuscular junction, and the mechanisms involved. Aging skeletal muscle is characterized by progressive muscle atrophy and a decline in contractile function, which negatively influences quality of life in elderly individuals and predisposes them to increased risk of falls, morbidity and mortality ,2. MusclConditions in which the neuromuscular junction undergoes degeneration precipitate severe muscle atrophy. In these conditions, retraction of motoneuron terminals with or without death of the entire motoneuron causes denervation of individual neuromuscular junctions. Denervation occurs cyclically, where a given denervation event is usually quickly followed by reinnervation by the original motoneuron (if it remains viable) or through collateral sprouting of an adjacent motoneuron axon . Importantly, severe muscle atrophy is temporally delayed from the initiation of the cycles of neuromuscular junction degeneration and is consequent to failure of reinnervation. Examples of diseases affecting the neuromuscular junction and which have rapidly evolving atrophy include amyotrophic lateral sclerosis (ALS) and the spinal muscular atrophies (SMA). Mouse models of ALS and SMA both show that neuromuscular junction alterations occur several weeks before onset of clinical symptoms ,12. At tAmong the earliest changes to be seen in aging muscle are alterations in neuromuscular junction morphology. Alteration of neuromuscular junction morphology was first reported in aged rodents in 1966  and was 1.5 (1.5 positive fibers (= denervated) are significantly atrophied versus young adult muscle fibers where no Nav1.5 positive fibers are seen. In contrast, Nav1.5 negative fibers in very old muscle are not significantly different in size from young adult  signaling, Cath aging . Althougth aging . In viewth aging  and functh aging ,31,32. Hth aging ,33, undeOur understanding of mitochondrial structure has evolved remarkably in recent years. Not only is the structure far more complex than the bean-shaped organelles that are still found depicted in some textbooks, but the structure is dynamic and tightly regulated. Perhaps nowhere else has this evolution in understanding of mitochondrial structure been more dramatic than in skeletal muscle. As early as 1978, the first indications that mitochondria likely formed a reticular-like structure in skeletal muscle were presented, based upon three-dimensional reconstruction of serial transverse electron micrograph images of rat diaphragm muscle . This waMitochondria in aged skeletal muscle have been reported to be both larger and more complex , and smaTo our knowledge, only one study has assessed mitochondrial morphology in aging motoneurons. This study found that mitochondria in motoneuron terminals at the neuromuscular junction in aged rats were swollen, had distorted cristae, and sometimes displayed evidence of membrane rupture  Figure . NotablyUCP1). These experiments showed that muscle-specific UCP1 over-expression induces alterations in neuromuscular junction morphology, including AChR fragmentation and degeneration of the motoneuron terminals [UCP2 and UCP3 are the major UCP isoforms in skeletal muscle and their role in mitochondrial uncoupling is less clear [Interrogating the functional consequences of alterations in mitochondrial structure is an evolving landscape, too. In the context of skeletal muscle, early studies making direct measures of mitochondrial function used mechanically isolated organelles . Howevererminals . Howeverss clear ,55. WhetAs noted above, mitochondria in motoneuron terminals of aged rats exhibit morphological features indicative of having undergone permeability transition . ConsistIn looking to explain why alterations in mitochondria occur with aging, multiple facets of mitochondrial biology need to be considered, including the fact that they have their own genome, that their structure is dynamically regulated (fusion and fission), that they are transported long distances in motoneurons to the terminals at the neuromuscular junction, and that they undergo continual turnover involving degradation (mitophagy) and replacement  to ensure high fidelity of organelle function. As such, understanding the basis for mitochondrial alterations with aging requires consideration of how aging impacts mtDNA (mutation and copy number), regulation of mitochondrial dynamics, and the mechanisms governing mitochondrial quality control or mitostasis. A summary of cited papers addressing mechanisms that may contribute to impaired mitochondrial function in muscle versus neurons with aging is provided in Mitochondria contain their own genome, the 16.6 kp long mitochondrial DNA (mtDNA), which consists of 37 genes encoding 13 polypeptides of the respiratory chain complexes, 2 rRNAs and 22 tRNAs . The remA special feature of mitochondria is the presence of multiple genomes per mitochondrion, such that when a mutation is present, both mutant and wild-type mtDNA copies co-exist in a single cell, in multiple cells of a given tissue, and/or in different tissues of a given individual. This phenomenon is known as heteroplasmy and it dictates whether normal mitochondrial function can be maintained . In factPolG) exhibits spontaneous mtDNA deletions and depletion, leading to an early appearance of traits that resemble premature aging [PolG mutant mouse study (fast twitch dominant) exhibit higher (not lower) levels of protein constituents of complex I, III and IV in very advanced age when atrophy becomes severe [PolG mutant mice suggest that high levels of mtDNA mutations and/or mtDNA depletion lead to different mitochondrial phenotypes than occur with normal aging at ages associated with severe muscle atrophy. This speaks to the fact that no prior studies have directly compared the nature of the muscle atrophy seen with normal aging to that occurring in the PolG mutant mouse to determine if mtDNA mutations and/or depletion phenocopies known aging muscle attributes. Such an analysis should include determining whether the PolG mutant mouse also exhibits neuromuscular junction degeneration, motor unit remodeling, and a failed reinnervation phenotype as seen with aging. These are areas that warrant further investigation to more fully understand the potential for mtDNA alterations to contribute to aging muscle deterioration.In contrast to the relatively uninspiring data from naturally occurring aging, a mouse model exhibiting a mutation in the proof-reading activity of polymerase gamma  has been studied in the context of age-related diseases such as PD; however, the role of mtDNA in motoneurons in the context of aging muscle atrophy remains under-studied. To our knowledge, there is only one study relevant to this point, and it was performed in post-mortem samples of spinal cord motoneurons of 14 elderly individuals (68–99 years of age) . This stMfn1 and Mfn2); and (ii) fusion of the inner mitochondrial membrane by optic atrophy 1 (Opa1). On the opposing side, mitochondrial fission is the process whereby a mitochondrion divides into two daughter mitochondria. Fission occurs under conditions of elevated stress and cell death, resulting in mitochondrial network fragmentation, a morphological state associated with mitochondrial dysfunction [Drp1), which forms a constrictive ‘necklace’ around regions of the mitochondrial network that tightens until two daughter mitochondria are formed. Fission also involves mitochondrial fission factor (Mff) and fission protein 1 (Fis1), which recruit Drp1 in the mitochondrial membrane [Opa1 [Mfn2 [Drp1 causes premature lethality  associated with muscle fiber loss, muscle fiber atrophy, and a more fused mitochondrial network [Opa1 and the fission protein Drp1 leads to various indices of abnormal mitochondrial morphology  and inhibits removal of dysfunctional mitochondria by mitophagy, which is accompanied by muscle atrophy and weakness [Opa1/Drp1 double knockout mice also exhibited indices of muscle denervation that resolved over time, suggesting that impaired mitochondrial dynamics in muscle alone can induce neuromuscular junction instability [Mfn2 cause Charcot-Marie-Tooth type 2A (CMT2A), an autosomal dominant disorder characterized by loss of motoneurons resulting in muscle atrophy [Mitochondrial dynamics govern mitochondrial number, size, morphology and function through the exchange of mitochondrial content  to meet variable energy demands, facilitate mitochondrial quality control, and spatially distribute mitochondria within a cell . This isfunction . Portionfunction   are associated with neurodegeneration, and deletion of m-AAA protease increases susceptibility to mitochondrial permeability transition [m-AAA proteases could play a role in the accumulation of mitochondria having undergone permeability transition in the motoneuron terminals of aged rats [YME1L, which not only degrades damaged or non-assembled proteins in the mitochondrial inner membrane but also cleaves Opa1 to activate it and catalyze inner membrane fusion [The importance of selective replacement of individual mitochondrial proteins is underscored by the remarkable heterogeneity in half-lives of different mitochondrial proteins . Althougansition . Thus, aged rats . Similare fusion , causes e fusion . The potPink1 and the E3 ubiquitin ligase, Parkin. Pink1 is transcribed in the nucleus, translated in the cytosol, and imported into the mitochondria through transporter of outer membrane (TOM) and transporter of inner membrane (TIM) complexes, ultimately localizing on the outer mitochondrial membrane of mitochondria. Although Pink1 is normally unstable due to high degradation rates, when mitochondria lose their membrane potential, Pink1 stabilizes on the OMM and initiates mitophagy by recruiting Parkin. Whereas the importance of both Pink1 and Parkin to mitophagy is well-established in neurons in general [Pink1 stabilization in skeletal muscle [Pink1 and Parkin causes motoneuron axon swelling and post-synaptic AChR cluster fragmentation at the neuromuscular junction degeneration [Parkin in neurons [Pink1 or Parkin prevents the age-related increase in mitophagy and exacerbates mitochondrial dysfunction [Parkin protein in aged human limb muscle. As alluded to above, in addition to its role in mitophagy, Parkin, ubiquitinates mitofusins and other mitochondrial proteins, and is thus involved in regulating mitochondrial network fragmentation and autophagic turnover of mitochondrial proteins [As noted above, mitophagy refers to the bulk degradation of dysfunctional mitochondria. The classical view of mitophagy involves a signaling pathway that includes the serine/threonine  general ,95, and l muscle . This nuneration ; changesneration , sensitineration  and causneration . It is aneration . Finally neurons , knockoufunction . Thus, ifunction  (Figure proteins . Parkin proteins . As suchAs stated above, biosynthesis of mitochondria requires the coordinated expression of both nuclear DNA- and mtDNA-encoded genes. The classical view is that mitochondrial biogenesis must, therefore, occur in close proximity to a nucleus. Whereas the multi-nucleated nature of skeletal muscle fibers provides for recurring so-called ‘nuclear domains’ along their length , motoneuThere is strong evidence for alterations in mitochondria with aging in both muscle fibers and motoneurons. Whereas there is experimental evidence that muscle-specific impairment of mitochondria can cause neuromuscular junction degeneration, it is not yet clear whether this occurs with aging. Indeed, there is evidence that some of the mitochondrial alterations seen in very advanced age in muscle are a consequence rather than a cause of denervation of muscle fibers. On the other hand, there is evidence that mitochondria in the terminals of aged motoneurons have undergone permeability transition and that this likely causes motoneuron die-back through retrograde transport of pro-apoptotic proteins to the motoneuron soma. As summarized in


Cellular reprogramming is a fundamental topic in the research of stem cells and molecular biology. It is widely investigated and its understanding is crucial for learning about different aspects of development such as cell proliferation, determination of cell fate and stem cell renewal. Other factors involved during development include hypoxia and epigenetics, which play major roles in the development of tissues and organs. This review will discuss the involvement of hypoxia and epigenetics in the regulation of cellular reprogramming and how interplay between each factor can contribute to different cellular functions as well as tissue regeneration. When Takahashi and Yamanaka were able to reprogram a fully differentiated skin cell into an induced pluripotent stem cell (iPSC), the promising potential of cellular reprogramming became evident. Through activation of critical transcription factors Oct4, Sox2, Klf4, and c-Myc (Yamanaka factors), any somatic cell could be reprogrammed into a pluripotent stem cell  or hypoxia (5% O2). Not only did the hypoxia treated cells grow faster, production of iPSCs significantly increased when cells were cultured under 5% O2 at all periods of induction  preferentially resulted in the maintenance of a highly proliferative, pluripotent population of human ESCs; Moreover, the culture at atmospheric 20% O2 levels expressed significantly less OCT4, SOX2, and NANOG than those maintained at 5% O2 in the hESCs , both mouse embryonic fibroblasts and human dermal fibroblasts were transduced with Yamanaka factors and cultured under normoxia . HIFs are activated in response certain environmental conditions, including a low oxygen environment and in inflammation , enzymes called prolyl-hydroxylases (PHDs) hydroxylate HIF-1α at proline residues 402 and 564. This modification tags HIF-1α for ubiquitination by the E3 ubiquitin ligase complex, which in turn tags it for degradation by the proteasome  renewal through its transcription factor HIF-1α and the Notch pathways . In cancers, the rapid outgrowth of cells and their consumption of oxygen overruns the limited supply of oxygen and creates a hypoxic environment in many areas of the tumor  muscle in mice after a laceration injury disrupted the vascular structure , results indicate that hypoxia could induce senescence of MSCs via altered gut microbiota  subunit and an RNA component. Telomeres and TERT are essentially in iPSCs induction and maintenance (Teichroeb et al., While the great majority of this review is focused on histone methylation, the potential role of histone acetylation and cellular reprogramming should not be ignored. An example of the impact of acetylation is the rapid acetylation that occurs after fertilization with the somatic nucleus. Before conception, the histones of mouse oocytes do not display any acetylation (Kim et al., Hypoxia has been proven to modulate histone acetylation by increasing histone acetylation at H3 and regulating the downstream processes of many genes. In addition, the initiation of hypoxia and induction of HIF-1α can increase the acetylation of H3 and H4 in neuroblastoma cells and increases the aggressiveness of neuroblastomas (Poljakova et al., Hypoxia stimulation greatly affects gene expression and is considered to play an essential role in early embryo development, cell differentiation, and cellular reprogramming. Meanwhile, new techniques have been introduced to this field, such as the development of next-generation sequencing, single cell RNA sequencing, and micro-omics technologies. Recent studies have clearly depicted certain epigenetic changes, including DNA methylation, hydroxymethylation, histone modifications, organoids and 3D structure formation during early embryo development, iPSCs, and somatic cell reprogramming. There is potential for the use of the interplay of hypoxia and epigenetics in regenerative medicine. The existence of reprogrammed cells after hypoxia has been reported in multiple different types of tissue and organs (Shyh-Chang et al., NN drafted the manuscript and created the figures. XS discussed and edited the manuscript. RD revised and edited the manuscript. YL searched literature, structured, modified figures, and wrote and edited the manuscript. All author approved the final version of the manuscript.YL was one of the executive editors for Journal of Cellular BioChemistry. The remaining 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.