Envisioning a T cell-dependent mechanism, they suggest that stimulation of sebum production by thymic stromal lymphopeietin (TSLP) “pulls” lipids out of the adipose tissue.The World Population Review (worldpopulationreview.com/) ranks obesity rates to one third of the global population. Associated co-morbidities comprise diabetes type 2, ischemic stroke and cardiovascular complications render this epidemic disease the number 1 cause of deaths worldwide (who.int/). Treating obesity encompasses besides classical approaches like dietary restrictions combined with increased physical activity more and more the targeting of molecular pathways involved in the pathogenesis and persistence of obesity. E.g., the discovery of “beige” adipocytes in white adipose tissue (WAT), which share lipid utilization via thermogenesis with “brown” adipocytes, fostered the hype to induce therapeutic “beiging” of WAT by cold stress and pharmacological interventions .1 In the NASH livers, TSLP also reduced hepatocyte damage, which is the consequence of lipid-induced oxidative stress and inflammation. TSLP might directly affect hepatocytes, since it stimulated hepatocyte autophagy after ischemia/reperfusion injury (I/R) by activation of the PI3-kinase pathway. This reduced liver damage in the I/R-induced sterile inflammatory environment,4 a scenario, which is closely linked also to the pathogenesis of the metabolic syndrome.While nutritional control by dietary or pharmacological interventions directly target the adipose tissue with the aim of fat tissue mass reduction, TSLP-stimulation of sebum production indirectly relates to the adipose tissue Fig. . The reg5 Yet, so far it is unclear, which form might be involved in sebocyte regulation in humans. Therefore, fat waste by TSLP-stimulated sebum production remains to be established unequivocally. Albeit the possibility to loose fat through the skin is intriguing, the stringent regulation of sebacious glands by hormones and neuropeptides and their feedback control requires careful evaluation before the clinical launch of TSLP as fat waist option. Hence, be aware of not casting out the Fiend with the devil of inflammatory skin and other organs´ diseases.Mouse and human TSLP are different. While in the mouse only one form exists, humans feature a long and a short isoform of TSLP. In mice, TSLP at physiological levels seems involved in homeostasis regulation, while overexpression supports adipose loss via increased sebum production. In humans, this relationship remains elusive. Current knowledge rather suggests that the long isoform is upregulated in Th2-mediated inflammatory allergic reactions like asthma and atopic dermatitis. The short form is constitutively expressed by, e.g., stimulation by commensal bacteria, and is involved in homeostatic regulation like maintenance of physiological gut barrier functions. The stimulation of epithelial cells by pathogens induces secretion of the long form.


PTBP3, ESRP1, and ESRP2 were demonstrated to contribute to position-specific AS. RNA-binding motif enrichment analysis near alternatively spliced exons provided further insight into the combinatorial regulation of AS at different positions by different RNA-binding proteins. These data, along with gene ontology (GO) analysis, represent a comprehensive analysis of the dynamic regulation of AS at different positions in chick cochlea.Alternative splicing (AS) refers to the production of multiple mRNA isoforms from a single gene due to alternative selection of exons or splice sites during pre-mRNA splicing. It is a primary mechanism of gene regulation in higher eukaryotes and significantly expands the functional complexity of eukaryotic organisms, contributing to animal development and disease. Recent studies have shown that AS also influences functional diversity by affecting the transcriptomic and proteomic profiles in a position-dependent manner in a single organ. The peripheral hearing organ, the cochlea, is organized to detect sounds at different frequencies depending on its location along the longitudinal axis. This unique functional configuration, the tonotopy, is known to be facilitated by differential gene expression along the cochlear duct. We profiled transcriptome-wide gene expression and AS changes that occur within the different positions of chick cochlea. These analyses revealed distinct gene expression profiles and AS, including a splicing program that is unique to tonotopy. Changes in the expression of splicing factors  Alternative splicing (AS) refers to the production of multiple mRNA isoforms from a single gene due to alternative selection of exons or splice sites during pre-mRNA splicing. It is a primary mechanism of gene regulation in higher eukaryotes and significantly expands the functional complexity of eukaryotic organisms. In humans, over 90% of multi-exon genes are alternatively spliced . It is wAuditory sensory organs such as the organ of Corti in mammals and the basilar papilla in avians are responsible for receiving and relaying sound information from the environment to the brain. These sensory organs reside within the elongated tubular structure known as the cochlea. Interestingly, auditory receptor cells, the hair cells, are specialized to detect different frequencies of sounds depending on their location along the cochlear duct. The hair cells located at the basal  regions of the cochlea are tuned to detect high frequencies of sounds, whereas the hair cells at the apical  cochlear regions are tuned to lower frequencies. This position-dependent frequency discrimination capability of the auditory system is known as the tonotopy. Hair cells display gradually different morphological features along the tonotopic axis, which may be crucial for discriminating various sound frequencies. Interestingly, some of the tonotopic features are conserved between the chicken and mouse. For example, the length of the hair bundle, which are composed of tens of stereocilium arranged in a stair-case pattern, is shorter at the base and gradually longer toward the apex, whereas the width of hair bundle is wider at the base and gradually narrower toward the apex .DNER, BMP7, and retinoic acid are also differentially expressed and regulate morphological features of hair cells important for discriminating sound frequencies in the developing chicken cochlea  of chick cochlea and performed an RNA-seq experiment  for both DEG and AS analyses results, confirming that the DEGs and AS events predicted from RNA-seq are true positives.RNA-binding protein (RBP) motif enrichment analysis near alternatively spliced exons showed motifs of multiple RBPs are enriched near alternatively spliced exons, providing further insight into the combinatorial regulation of AS at different positions by different RBPs. Gene ontology (GO) analysis using genes detected from DEG analysis and AS analysis revealed terms highly related to the developmental process and sound perception.2 inhalation followed by decapitation. The head was hemi-sectioned and the cochlea duct was isolated in cold DEPC-treated PBS. The basilar papilla was dissected from the cochlear duct and divided into three pieces: base, middle, and apex. Each piece was collected in a separate Eppendorf tube containing Trizol reagent  and quick-frozen to minimize RNA degradation. Total RNA from 12 chickens (24 cochleas) were used for 3 biological replicates for each region . RNA was isolated using Trizol reagent according to the manufacturer’s instructions.Post-hatch day 1 (P1) chickens were euthanized by COWe performed an RNA-seq experiment for three sample groups on the base, middle, and apex regions of chick cochlea. Following cochlea dissection and mRNA extraction, the quality and quantity of RNA were examined using Tapestation2100–RNA Screen Tape (Agilent) and a Victor3 Multilavel plate reader (PerkinElmer), respectively. 1μg of total RNA from each sample was reverse transcribed into cDNA and amplified using the Illumina TruSeq RNA Sample Prep Kit v2. For library preparation, mRNA was enriched by oligo-dT to minimize DNA contamination. The PCR products for the cDNA library were purified and selected using the AMPure XP bead (Beckman Coulter). The DNA was quantified using Tapestation2100–D1000 Screen Tape. The cDNA libraries were multiplexed and sequenced on the Illumina Hi-Seq 4,000 platform, and 101-bp paired-end reads were generated. The files from the multiplexed RNA-seq samples were demultiplexed, and fastq files for each sample were generated. Approximately 57–82 million read pairs per replicate were generated. To quantify the transcript abundance, sequencing reads were aligned to the chicken genome  and the transcriptome (Ensembl GTF release 91), using the STAR aligner (v2.5.4b) , allowin• Up-trend includes genes that show at least a 1.5-fold increase in mean FPKM values with directionality from base to middle to apex, allowing up to 15% variability against the directionality.• Down-trend represents genes that show at least a 1.5-fold decrease in mean FPKM values with directionality from base to middle to apex, allowing up to 15% variability against the directionality.We identified DEGs on base, middle, and apex regions using Cuffdiff (v2.2.1) . We calcp values and FDRs of splicing events with a cutoff |Δψ| of >0.01%, then significant splicing events were identified at FDR <5% and |Δψ| ≥ 5% as described previously  , which ceviously .To validate DEGs and AS events detected from RNA-seq data, we performed reverse transcription (RT)-PCR using extracted total RNA as described previously . Total RWe sought to identify binding sites of splicing factors and other RBPs that were significantly enriched in differential exon skipping events. To assess the enrichment of RBPs near alternatively spliced exons, the differentially spliced exons from rMATS runs were compiled. These differentially spliced exons were scanned for RNA binding protein motifs using rMAPS2  with defWe conducted two sets of GO analyses: the one with DEG results and the other with AS results. 1,479 Up-trend genes and 797 Down-trend genes from DEG analysis were compiled for the first set of GO analysis using PANTHER  with GO To evaluate phenotypes associated with different regions along the longitudinal axis of the cochlea duct, we extracted mRNA from three different regions of the chick cochlea, base, middle and apex, and performed an RNA-seq experiment. This yielded 57 to 82 million read pairs per replicate, which were then aligned to the chicken genome. The transcript abundance was quantified, and FPKM values for each sample were obtained. Of 24,880 Ensembl-annotated genes, there were 13,045 (52.4%) in which at least one of the 9 samples had an FPKM ≥1. To assess position-specific gene expression that might be responsible for configuration and maintenance of the tonotopic axis of the cochlea duct, we identified differentially expressed genes on the three regions. We compared gene expression profiles from two different regions of cochlea at a time for all three regions. Since gene expression profiles originate from the same cochlea tissue despite being extracted from different regions, they should show intrinsically much similar gene expression patterns compared to those from different organs. Thus, we searched for DEGs with more relaxed criteria. We considered a 1.5 fold-change in expression level as a significant difference along three different regions.To identify genes that are responsible for tonotopic configuration along the cochlea duct, we searched for genes that gradually increase their expression by a factor of at least 1.5 from base to middle to apex regions and designated them as Up-trend. Likewise, we searched genes that gradually decrease their expression at least 1.5-fold from base to middle to apex regions and designated them as Down-trend. For these gene expression gradients, we identified 1,479 Up-trend genes and 797 Down-trend genes. The gene expression patterns along the regions of the cochlea are represented in BMP7, DNER, KCNJ2, TECTB, and TMC2 , in which a cassette exon around two closely located exons is either included or skipped. From the three comparisons, we identified 275, 114, and 325 significant SE events, respectively . Of thesCDH23, EPB41L3, KCNMA1, and LMO7  and syndromic (USH1D) hearing loss in humans , and its alternative splicing has been suggested to be important for frequency tuning along the tonotopic axis by influencing the channel gating properties  pathway by introducing stop codons unexpectedly out of the original frame due to non-canonical AS events. As transcripts that have retained intron (RI) often contain premature stop codons, which can trigger the NMD , we searThese results suggest that differential AS events along the cochlear duct act as a fundamental mechanism for establishing the tonotopy by altering morphological and functional properties of the proteins essential for frequency tuning. Although these genes represent gene expression gradients in AS events, they are not necessarily directly related to gene expression gradients at the transcription level.We further evaluated the characteristics of the genes detected from DEG analysis with Gene Ontology (GO) enrichment analysis using PANTHER . For theWe next performed GO enrichment analysis on genes with differential skipped exon events using PANTHER  Figure . Enrichep-value) while the PTB like motifs were enriched in the upstream flanking intron regions when the inclusion of the target exon is suppressed . We also examined the enrichment of ESRP 1/2 motifs , we performed motif enrichment analysis using rMAPS2  with dif2 motifs . There wWe report that alternatively spliced transcript variants encoding different protein isoforms are closely related to the configuration along the tonotopic axis of the chicken cochlea as well as differentially expressed genes. Unlike DEGs on two different organs or two different treatments, DEGs on the different regions along the longitudinal axis of the cochlea show subtle changes on the expression level. By applying relaxed criteria, we found DEGs that represent gene expression gradients along the tonotopic axis, from base to middle to apex regions where sounds are detected from higher frequencies to lower frequencies. We performed RNA-Seq analysis and identified 1,479 Up-trend and 797 Down-trend genes that might be responsible for configuring the tonotopic axis and maintaining the cochlea’s function in detecting different frequencies of sound. Among DEGs, as expected, we identified genes that were responsible for regulating membrane proteins and ion channels that affect structural formation, signaling, and sensory function. We further validated our RNA-seq analysis results with RT-PCR for some of DEGs to confirm that the RNA-seq data set reflected actual gene expression at the transcription level.MYO7A, CLRN1, WHRN, FSCN2, PCDH15, PLS1, CIB2, USH1C, THRB, and PTPRQ. On the other hand, in Down-trend DEGs expressed higher levels at the base, enriched GO terms include broad developmental and biological processes such as system development, multicellular organismal processes, and signal transduction. GO terms enriched in molecular function and cellular component are also general biological terms but not specific to the inner ear. These results suggest that tonotopy-specific features are mainly dictated by Up-trend DEGs, while general developmental and biological processes are maintained by Down-trend DEGs.GO analysis using Up-trend and Down-trend DEGs shows that different GO terms are enriched in each group. In Up-trend DEGs expressed at higher levels in the apex, GO terms related to the “inner ear” are enriched. For example, GO terms among enriched biological processes include inner ear development, ear development, sensory perception of sound, and inner ear receptor cell development. In addition, enriched GO terms in molecular function and cellular component are mostly related to hair cell function and morphology such as ion channel activity and stereocilium. Consistent with these GO analysis results, Up-trend DEGs include many genes related hearing loss such as KCNMA1 gene, which was reported to have 7 different splicing sites  and Cdh23 (−68), are produced in the mouse hair cells. Our analysis show that this inner ear-specific AS events of CDH23 gene also occur in chicken. The chick CDH23 gene has 33 exons, and we found CDH23 (+32) and CDH23 (−32) splicing variants in chick basilar papilla. Interestingly, the amino acid sequences of the inclusion exon (exon 68 in mouse and exon 32 in chicken) show 96.4% similarity, suggesting that the inner ear-specific exon inclusion is conserved in avians and mammals. Moreover, our AS analysis further revealed that this AS event occur differentially along the tonotopic axis  ganglion neurons suggests its possible role in the regulation of neuronal gene expression and differentiation in the developing mouse cochlea (Srrm4, which encodes a splicing factor, causes hearing loss in mouse (We further identified regulatory RBPs, which may play essential roles in differential AS events along the tonotopic axis. We noted that PTB motif sequences are enriched around skipped exons in either the upstream intron region of the exon or in the exon and downstream intron regions. While functional validation of PTBP1 in the AS events along the cochlea warrants further investigation, it is noteworthy that the inclusion of exon 68 of by PTBP1 . In addi cochlea . Furtherin mouse . Since tin mouse We also found that the intensity of transcriptional expression of splicing factors does not necessarily correspond to the gene expression levels along the tonotopic axis, suggesting that there could be additional regulatory factors involved in AS such as signaling genes and ligand-binding proteins .Given the lists of genes from DEG and AS analyses, we identified biological processes that are represented with those genes and opened the possibility to find the functions of some genes whose roles are not yet fully understood in establishing the tonotopy. Since many morphological and functional features are conserved in avian and mammalian cochlea, DEGs and AS events identified in the chicken cochlea will greatly help exploring the mechanisms contributing to cochlear development and function as well as hearing loss in mammals such as mice and humans. Moreover, comparative analysis of DEGs and AS events between chicken and mouse cochlea may shed light on uncovering the mechanism of hair cell regeneration occurred specifically in chicken. In summary, the overall gene expression profiles, specifically AS events along the tonotopic axis of the chick cochlea, are orchestrated by complex gene expression mechanisms in addition to the differential expression of genes depending on different regions of the cochlea. We show that the detection of subtle frequency differences of sound throughout the longitudinal axis of the chicken cochlea is sophisticatedly regulated by differential expression of genes and that AS takes an important role in regulating gene expression gradients along the tonotopic axis.


Neuroendocrine tumors are a collection of neoplastic lesions arising in cells with traits similar to hormone-producing and nerve cells with the ability to secrete peptide hormones using an intricate vesicle transportation system. From a clinical standpoint, neuroendocrine tumors are unique in terms of therapeutic modalities, and a correct diagnosis is therefore imperative in order for the patient to obtain the most efficient treatment. In this process, the pathologist can analyze if the tumor cells express Chromogranin A and Synaptophysin, two proteins associated with the regulation of secretory vesicles. Unfortunately, these markers are not always present in neuroendocrine tumors, and non-neuroendocrine tumors may also occasionally express Chromogranin A or Synaptophysin—making the diagnosis difficult to make for certain cases. Recently, three proteins termed ISL1, INSM1 and Secretagogin were found to be selectively expressed in neuroendocrine cells, and subsequent studies have identified their potential as markers of neuroendocrine differentiation in the clinical setting. In this commentary, the benefits of these novel “second-generation” markers are briefly discussed from a clinical context. When analyzing tumors by histopathology, endocrine pathologists have traditionally been restricted to a few key immunohistochemical markers related to secretory vesicles in order to pinpoint neuroendocrine differentiation—most notably Chromogranin A (CGA) and Synaptophysin (SYP). Although proven of great clinical utility, these markers sometimes exhibit tissue-specific patterns depending on tumor origin, and non-neuroendocrine tumors might sometimes display focal expression. Moreover, CGA and SYP might be partially or totally absent in highly proliferative neuroendocrine carcinomas, making the diagnosis particularly challenging on small biopsies of metastatic lesions with unknown location of the primary tumor. The advent of second-generation neuroendocrine markers ISL LIM Homeobox 1 (ISL1), INSM Transcriptional Repressor 1 (INSM1) and Secretagogin (SECG) have expanded the pathology toolbox considerably, constituting markers that often retain expression even in poorly differentiated neuroendocrine carcinomas. As non-neuroendocrine tumors seldom express these antigens, the specificity of ISL1, INSM1 and SECG make them welcome additions to clinical practice. In this commentary, recent advances of this field as well as initial clinical experiences from a tertiary neuroendocrine center are discussed. Although pathology laboratories are diverse in terms of the amount of immunohistchemical markers available for diagnostic purposes, the most commonly employed markers in the context of diagnosing a neuroendocrine tumor (NET) constitute Chromogranin A (CGA), Synaptophysin (SYP), Cluster of differentiation 56 (CD56) and Neuron-specific enolase (NSE). CGA is essential for the maturation secretory granules in many endocrine cells, and also regulate the exocytosis process . Its useISL LIM Homeobox 1  was originally found to be selectively expressed in neuroendocrine and neuronal cells, and has since been established as a transcription factor with important roles orchestrating pancreatic endocrine differentiation ,13. SpecIn all, these three proteins are functionally distinct from CGA and SYP, and have multiple roles unrelated to the vesicle transportation system. From a biological context, these markers may therefore be consistent in terms of expression even if the NET/NEC downregulates its secretory machinery as part of the dedifferentiation process. In endocrine pathology, immunohistochemical markers are well-attributed for their diagnostic and prognostic features, not least in terms of neuroendocrine neoplasia ,30,31,32Yet another clinical issue is the proper identification of poorly differentiated NECs with high proliferation counts, as these tumors not seldom show reduced expression of CGA and SYP . ConsultAn additional aspect of diagnosing metastatic NETs/NECs with unknown primaries is the fact that tissue-based expressional patterns not always are consistent. Pathologists may use CK7/CK20 as blunt tools followed by combinations of transcription factors PDX1, Caudal Type Homeobox 2 (CDX2), Thyroid Transcription Factor 1 (TTF1) and Paired Box 8 (PAX8) to identify the tissue of origin ,38, but bona fide NET component as well as a non-NET population  are visualized [The complexity of endocrine pathology is perhaps best exemplified by the NET/non-NET mixed tumor entity entitled “MiNEN”, in which a sualized ,41. In tDespite the rapid development of next-generation sequencing techniques, immunohistochemistry still enjoys the top position as the most recurrently used adjunct histopathological methodology in clinical routine. Fast, cheap and easy to interpret when standardized, the method is standard practice in modern pathology laboratories. Visualizing immunohistochemical markers in cancer diagnostics is a clinical cornerstone with great diagnostic and prognostic utility, and expressional patterns might also guide oncologists to specific therapeutic alternatives. Moreover, information regarding the immunoreactivity of a specific protein could also triage patients for further genetic counseling. When diagnosing NETs, the pathologist is largely dependent on this technique to safely visualize neuroendocrine differentiation, as morphology alone is usually not sufficient to properly diagnose a tumor as “neuroendocrine”. Traditionally, first-generation markers such as CGA and SYP have been widely used and reproduced, and are still considered gold standard in endocrine pathology. However, non-NET mimics with focal neuroendocrine differentiation are not entirely uncommon, and highly proliferative NECs recurrently lose the ability to express CGA and SYP. Moreover, CGA and SYP offer little help to the diagnostician in terms of a tumor’s primary location if this is not known. To counter this, the inclusion of second-generation neuroendocrine markers ISL1, INSM1 and SECG could (1) increase the likelihood of correctly pinpoint NETs when a non-NET is among the potential differential diagnoses, (2) identify poorly differentiated pancreatic and pulmonary NECs exhibiting absent/focal CGA and SYP immunoreactivity and also (3) give important clues regarding the origin of metastatic cases when a primary tumor has not been established through radiology and/or clinical examinations. Pathologists in tertiary centers with high NET volumes are encouraged to establish second-generation neuroendocrine markers as part of the clinical routine arsenal in order to improve the diagnostic capability.


Secundum-type atrial septal defects (ASD) constitute 8% to 10% of congenital heart defect. Secundum ASDs can be closed either percutaneously or surgically. However, ASD device closure has proven to be technically safe and feasible; it is not free of complications. These complications include device embolization/malposition which have been reported in 3.5% of cases, arrhythmia, and pericardial effusion in 2.6% and 0.5–1.5% respectively, device thrombus, residual shunting, and impingement of the device on the adjacent structures.We introduce three patients with secundum ASD who had preexisting pericardial effusion, device closure was performed for them, and after the procedure, the effusion size progressed significantly. We used multimodality imaging to diagnose the cause of pericardial effusion (PE). Cardiac erosion was diagnosed in one of the patients that managed surgically. We did not found any specific procedure-related cause for worsening the pericardial effusion in the other two patients.Several reasons include procedure-related complication and other systemic causes should be considered in patients who develop pericardial effusion after trans catheter closure of ASDs. Secundum-type atrial septal defects (ASDs) constitute 8% to 10% of congenital heart defect [Trans catheter closure (TCC) has dramatically improved the management of ASDs in such a way that TCC is being the method of choice for closing the defects in eligible ones . ASD devPericardial effusion after device closure is one of the ASD closure complication that could be resolved spontaneously, but it might be a sign of a catastrophic complication; it means cardiac erosion. Erosions in adjacent cardiac structures are very rare (0.2–0.3% of cases) after TCC of ASDs and in most cases happen within the first 6 months after ASD occluder implantation .The true diagnosis for an appropriate management is a prime issue in those with post TCC of ASDs with pericardial effusion. By the way, we also have cases with preexisting PE whom the amount of the effusion has gotten worse after the procedure.In this paper we introduce three patients with secundum ASD who had preexisting pericardial effusion and device closure was performed for them and after the procedure, the effusion size progressed significantly. We discuss about the individualized approach and management of these patients.A 55-year-old female patient, a known case of end-stage renal disease (ESRD) on hemodialysis, was referred to our hospital after being diagnosed with a 20-mm secundum ASD in pre-operative evaluation for renal transplantation. In transesophageal echocardiography (TEE), moderate right ventricle enlargement and pulmonary hypertension were detected. Pre-procedural TEE showed: Normal left ventricle (LV) systolic function (LVEF: 55%) with LV diastolic dysfunction grade I, moderate right ventricle (RV) enlargement, and mild systolic dysfunction. A large secundum ASD with a significant left to right shunt with sufficient rims for device closure was notable. Moderate tricuspid regurgitation (TR) with tricuspid regurgitant gradient (TRG), 40 mmHg, and moderate pulmonary arterial hypertension (PAH) were reported. Mild circumferential pericardial effusion (PE) was also demonstrated Fig. . The patTTE on the first day after the procedure revealed moderate to severe PE Fig. . Spiral A 34-year-old woman presented to our hospital with complaints of exertional dyspnea  who in TTE had mild to moderate RV enlargement and dysfunction with ASD secundum and mild to moderate circumferential PE , Spodick et al. found 13 of their 40 patients (32.5%); > 21 years of age with isolated secundum ASDs have circumferential PE preoperatively [Post ASD closure pericardial effusion occurs with a reported incidence of 0.5–1.5 % secondary to cardiac perforation, device erosion, or reactive PE. Pre-existing PE could be due to different causes include uremic, autoimmune, hypothyroidism, Viral, bacterial, fungal or parasitic infections. In a retrospective study  which in this patient confirmed the disastrous complication, it means cardiac erosion; so, immediate surgical management of the patient was performed.The second patient had mild to moderate pre-existing PE with no response to anti-inflammatory agents during 2- to 3-month follow-up that seems to be due to ASD itself. With mild to moderate pre-existing PE, Reddy et al. speculated that, if septal defects were left untreated, the effusion may accumulate over time because of chronic volume overload, which may lead to more PE . The amoThe third patient had mild to moderate PE due to unknown etiology that after device implantation increased to moderate PE without evidence of cardiac perforation, erosion, or hematoma, so we managed her conservatively. Fortunately, she was asymptomatic and the PE decreased significantly within 3-month follow-up.Generally speaking, several reasons should be considered in patients who develop pericardial effusion after TCC of ASDs; the most and catastrophic cause is cardiac erosion which frequently happens at the left atrium roof or aortic side. So, the immediate diagnostic modality which could rule out is cardiac CTA; if no evidence of cardiac erosion was found, looking for hematoma around the pulmonary vein or left atrium appendage which commonly occurs by wire manipulation which leads to perforation should be appraised. In fact, the diagnosis of cardiac erosion was confirmed by detection of hemopericardium based on the density and Hounsfield CT number of the pericardial fluid and retroaortic hematoma in the first patient. Echocardiography may not be enough to rule out cardiac erosion. CTA can reveal information about structures that are difficult to visualize with TEE such as erosion of an ASD closure device into adjacent structures and performing cardiac CTA and looking for the evidence of cardiac erosion are essential .The uncommon cause of PE, post TCC of ASD is case reports of seroritis (pleuro-pericarditis) which the underlying stimulus is not clear, but one hypothesis is the reaction to Nickle and the materials of the device; it seems to respond well to anti-inflammatory agents with low recurrence rate. In some patients, the underlying advanced RV failure leads to persistent PE even after ASD closure who have a very poor prognosis. In fact, this group of patients with advanced right ventricular failure which is somewhat irreversible develop pericardial effusion after ASD closure due to RV failure and require treatment for right ventricular failure and they have increased risk for atrial and ventricular arrhythmia. finally, in some cases, no reasonable source of PE could be found and extensive work-up for acute, sub-acute, and chronic pericarditis should be performed.Our experience in the management of these three patients highlighted that recognizing the different causes of pre-procedure and post-procedure pericardial effusion using diagnostic techniques such as echocardiography as well as another imaging modality such as cardiac CTA could be very useful and important for proper diagnosis, and management of worsening pre-existing PE in patients after ASD device closure. Cardiac CTA is a reliable diagnostic method for the detection of device erosion or cardiac perforation in patients who underwent transcatheter ASD closure with new onset or worsening of pre-existing PE.


Background: Neurocardiac dysfunction worsens clinical outcome and increases mortality in stroke survivors. We hypothesized that heart rate variability (HRV) biofeedback improves neurocardiac function by modulating autonomic nervous system activity after acute ischaemic stroke (AIS).Methods: We randomly allocated (1:1) 48 acute ischaemic stroke patients to receive nine sessions of HRV- or sham biofeedback over 3 days in addition to comprehensive stroke unit care. Before and after the intervention patients were evaluated for HRV via standard deviation of normal-to-normal intervals , root mean square of successive differences between normal heartbeats (RMSSD), a predominantly parasympathetic measure, and for sympathetic vasomotor and sudomotor function. Severity of autonomic symptoms was assessed via survey of autonomic symptom scale total impact score (TIS) at baseline and after 3 months.Results: We included 48 patients with acute ischaemic stroke . Treatment with HRV biofeedback increased HRV post intervention  and alleviated autonomic symptoms after 3 months , which was not seen after sham biofeedback . There were no changes in sympathetic vasomotor and sudomotor function (p = ns).Conclusions: Adding HRV biofeedback to standard stroke unit care led to improved neurocardiac function and sustained alleviation of autonomic symptoms after acute ischaemic stroke, which was likely mediated by a predominantly parasympathetic mechanism.Clinical Trial Registration:www.ClinicalTrials.gov, identifier: NCT03865225. To avoid confounding of autonomic function assessment we excluded patients with history or clinical evidence of autonomic neuropathy, intake of any tricyclic antidepressant within the last 14 days or atrial fibrillation. Furthermore, we excluded patients who were considered incapable of performing biofeedback because of aphasia, cognitive deficits, respirational insufficiency, blindness, deafness, malignant cerebral infarct, indication for treatment at intensive care unit or other physical limitations with resulting inability to participate in HRV biofeedback training.randomizer.org) and used sequentially numbered containers to conceal the sequence until interventions were assigned. Recruitment of study participants was undertaken from November 2018 to August 2019. At baseline all patients underwent medical history assessment and physical examination including evaluation for neurological deficits as well as autonomic testing of neurocardiac, sudomotor and vasomotor function. The study intervention, HRV biofeedback, was performed over 10 min, three times a day for three consecutive days, starting at baseline in a single masked setting where patients were blinded to the group allocation (sham control vs. biofeedback). Autonomic testing was repeated immediately after the last biofeedback training. Assessment of severity of autonomic symptoms and functional impairment was performed at baseline and was repeated by means of a phone-based follow-up after 3 months. All autonomic and clinical assessments were performed by an investigator who was not blinded to treatment allocation (P.O.). Statistical analysis was performed by an investigator (K.B.) blinded to group allocation.We randomly allocated 48 patients with acute ischaemic stroke (1:1) to receive nine 10-min sessions of either HRV biofeedback or sham biofeedback over 3 days in addition to standard stroke unit care. An investigator (P.O.) generated the allocation sequence using an online randomizer  with associated ear pulse sensor was used for HRV biofeedback training with continuous assessment and visualization of HRV on the computer screen. In this visualization, a butterfly was hovering in the air and was ascending when HRV increased or descending when HRV decreased, respectively. Breathing instructions were given on the screen as a moving bar with upward movement signaling the patient to inhale and downward movement indicating to exhale. After having completed a test-training, patients underwent nine 10-min biofeedback training sessions over a period of 3 days. Frequency and duration of the intervention was determined in consideration of compatibility with standardized care procedures as part of stroke unit care as well as previously published intervention protocols and explorative studies of short-term HRV biofeedback  and were converted using a four-channel-digitizing polygraph . Analyses were carried out using the software package LabChart® for Windows . In order not to interfere with stroke unit care all measurements took place in the patient's room of our stroke unit with a controlled room temperature of 21–25°C. Autonomic testing was performed in a semi-recumbent position after a 10-min lasting rest.All biosignals derived from autonomic testing of neurocardiac, sudomotor and vasomotor function signals were processed by an artifact-filter and signal amplifier  recorded cardiac electrical activity over two phases of 3 min each. In the first phase, patients were instructed to breathe casually, whereas in the second phase patients breathed at a fixed frequency of six per minute with an inspiration/expiration ratio of 1.5/1 as indicated by an audio signal to increase parasympathetic activity and thereby HRV . The maximum increase in amplitude following was calculated as a measure of the sympathetic sudomotor response.The sympathetic skin response was measured to assess sudomotor function as previously desribed . BrieflyCutaneous blood flow following sympathetic stimulation was measured using a plethysmograph  equipped with an infrared photoelectric sensor as previously described . BrieflySeverity of symptoms attributed to the autonomic nervous system were assessed using the German translation of the Survey of Autonomic Symptoms , 15. Thet-test for independent samples, Wilcoxon rank-sum test, Chi-square test and Fisher's exact test were applied to compare baseline characteristics between groups, where applicable. The primary outcome was change of SDNN from baseline to post intervention under paced breathing. Further outcomes of clinical interest comprised RMSSD, spectral analysis parameters of HRV , parameters of autonomic sudomotor (sympathetic skin response) and vasomotor  function, and autonomic symptom severity (TIS). Since autonomic data followed a non-normal distribution, the Wilcoxon signed-rank test was used for dependent within-subject comparisons and the Wilcoxon rank-sum test for independent between-subject comparisons. Data are expressed as median [interquartile range], mean (± standard deviation) or percentage according to type and distribution.All statistical analyses were performed with STATA software . Following testing for normality and equality of variances, Student's t-test for independent samples, Wilcoxon rank-sum test, Chi-square test, Fisher's exact test, Wilcoxon signed-rank test or McNemar's chi-square test were applied where applicable in the intention-to-treat population to compare outcomes with statistical significance set as p-value <0.05. While the datasets of our primary outcome and all autonomic tests were complete, missing data points in the 3-month follow up were considered missing at random and were treated using an available case analysis. All statistical analyses were performed by original assigned groups.Due to the exploratory nature of this study no formal sample size calculation was undertaken. Sample size of this pilot study was determined based on a previous study of HRV biofeedback in patients with alcohol dependence showing cardiac autonomic improvement post intervention . Studentn = 24) did not differ from those in the sham-biofeedback group (n = 24) with respect to sex, age, body mass index, alcohol consumption, cardiovascular comorbidities, medication, clinical baseline value, stroke etiology and acute treatment , median (interquartile range)] with acute ischaemic stroke. The study was ended after complete recruitment of the predetermined sample size. All analyses were by original assigned groups. Patients in the HRV biofeedback group  displayed an increase in HRV under paced breathing condition on time-domain analysis with elevated SDNN (n = 24). The predominantly parasympathetic HRV measure RMSSD was also elevated following HRV biofeedback but not sham biofeedback  baseline vs. 40.2 (30.8) ms post-intervention, p = 0.63; sham biofeedback, n = 24: 25.4 (28.2) ms baseline vs. 29.5 (33.5) post-intervention p = 0.46] nor via RMSSD .There was neither such change when cardiac autonomic function was assessed during normal breathing Furthermore, analysis of spectral components of HRV under paced breathing conditions showed an increase in low frequency band and total power in patients who underwent HRV biofeedback but not in those who received the sham intervention.In contrast, there were no such changes when spectral analysis of HRV was undertaken under normal breathing .Assessment of sympathetic skin response neither revealed any changes of sudomotor autonomic function in patients with acute ischaemic stroke following HRV biofeedback training nor in those who have undergone sham biofeedback . Adding p = 0.029; sham biofeedback: 4.0 (7.0) vs. 5.5 (7.5), p = 0.06]. As expected functional deficits measured after the intervention were decreased in both study groups .The addition of HRV biofeedback to standard stroke unit care led to an alleviation of severity of autonomic symptoms 3 months post intervention, which was not seen after sham biofeedback [HRV biofeedback 3.5 (8.0) vs. 7.5 (7.0) baseline, The major finding of this randomized sham-controlled pilot study is that HRV biofeedback might improve neurocardiac function and alleviate autonomic symptoms in patients with acute ischaemic stroke undergoing stroke unit care. The treatment was well-tolerated by stroke patients and its integration into the multidisciplinary setting of a comprehensive stroke unit was feasible. Improvement in parasympathetic function following HRV biofeedback was neither paralleled by improvement in measures of sympathetic function nor in sympathovagal balance, indicating a predominantly parasympathetic mechanism of action. Lastly, we were able to gather pilot data to provide a basis for calculating the sample size of follow-up research to confirm these findings and study the interventions' effects on functional recovery from acute ischaemic stroke.Multiple clinical studies have shown that HRV biofeedback can alleviate neurocardiac dysfunction and improve clinical outcomes in neuropsychiatric and cardiovascular disorders, possibly mediated by augmented respiratory sinus arrhythmia triggering increased baroreflex gain and parasympathetic outflow , 21. ThiSpectral analysis of HRV showed an increase of total power after HRV biofeedback, consistent with an overall increase in HRV. Analysis also revealed an increase in low frequency, a HRV component considered to be determined by both branches of the autonomic nervous system. This could indicate a sympathetic component to the mechanism whereby the study intervention improved neurocardiac function but is in contrast with the absence of any changes in neurocardiac sympathovagal balance and non-cardiac sympathetic measures in our study. Alternatively, viewed in conjunction with the observed increase in SDNN and RMSSD after HRV biofeedback, these findings might support the previously reported superior precision of time domain over spectral analysis of frequency domains in the evaluation of neurocardiac function . This isFeasibility of implementing HRV biofeedback into a standardized multidisciplinary stroke unit care as well as high tolerability of the treatment procedure in our study indicate that the treatment can be added to standard care of patients with acute ischaemic stroke to reduce neurocardiac dysfunction post-stroke without jeopardizing integrative care. A previous randomized study of HRV biofeedback enrolled patients who have experienced an ischemic stroke within 1 week from stroke onset also showed an increase in HRV after treatment compared to a control group that received no study intervention . HoweverAlthough absolute differences in time domain measures of HRV were relatively small between patients treated with HRV biofeedback and sham control patients, they might be clinically meaningful since decreased HRV has shown to predict all cause-death and cardiovascular events with likely implication to cerebrovascular disease –32. HoweStrengths of our study include its randomized control design and the assessment of non-cardiac vasomotor and sudomotor sympathetic function measures allowing detailed phenotyping of autonomic functional integrity. We were able to test the implementation of HRV biofeedback treatment within a state-of the art comprehensive stroke unit, underscoring the probable external validity of our observations. Our study is limited by the lack of a long-term follow up assessment of neurocardiac function. Therefore, we do not know if the observed increase of HRV after treatment would be sustained in later phases of rehabilitative therapy. However our results suggest alleviation of autonomic symptoms after HRV biofeedback, which was found to be sustained 3 months post-intervention, indicating a possible beneficial long-term effect on functional integrity the autonomic nervous system. We did not perform any analysis on possible effects of onset-to-randomization time or ischaemic damage to specific autonomic control centers on treatment effects because of the limited sized of our study population. However these associations will be investigated in a larger confirmatory study. Moreover, our study provides pilot data for a follow-up trial in a larger study population, which will also include assessment of the treatment's efficacy in facilitating functional recovery from stroke. In contrast to the majority of previous studies on HRV biofeedback, we included a sham control to minimize placebo effect, a strategy that was shown to increase effect sizes compared to inactive control conditions .In conclusion, HRV biofeedback might be a feasible and efficacious tool to counteract neurocardiac dysfunction and alleviate autonomic symptoms following acute ischaemic stroke. However, broad clinical implementation of the treatment requires follow up research in a large study population of patients with acute ischaemic stroke to understand whether modulation of autonomic neurocardiac function may translate into improved functional recovery and prevention of recurrent cerebrovascular events.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.Clinicaltrials.gov registration: NCT03865225.The study was approved by the ethical review committee of Technical University of Dresden . Written and oral informed consent was obtained from each study participant. TS: supervision of data acquisition, drafting of the manuscript, study concept and design, analysis and interpretation of data. PO: acquisition of data (major role), revising manuscript for content, and interpretation of data. AS and JB: revising manuscript for content and interpretation of data. ES, MA, and L-PP: acquisition of data (supporting role) and revising manuscript for content. GR: analysis and interpretation of data and revising manuscript for content. HR and VP: study concept and revising manuscript for content. KB: study concept, analysis of data, and revising manuscript for content. 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.


In atypical hemolytic-uremic syndrome (aHUS), various defects of the complement system have been reported to explain pathophysiology. Therapeutic options for complement inhibition are well-recognized; however, the links between various immune-derived diseases and aHUS are unclear, and their interference with treatment efficacy during long-term complement-blocking therapy is scarcely known.We present a pediatric patient who developed aHUS with acute kidney injury in parallel with the onset of Crohn’s disease (CD), and who required long-term complement-blocking therapy with eculizumab (ECU). Unexpectedly, during the 6-year ECU treatment, an important intra-patient variation of the degree of complement inhibition was observed. In spite of continuous and stable doses of complement-blocking therapy, periods of incomplete blockade were observed in strong association with relapses of CD. When conventional and later biological therapy with adalimumab was introduced, with CD going into remission, complement blockade became complete again. Despite periodically low ECU levels and insufficient complement inhibition, no clinical or hematological signs of aHUS recurrence were detected during CD relapses.In aHUS cases secondary to CD, close monitoring of both complement inhibition and serum ECU levels is needed as intestinal disease can interfere with complement-blocking treatment. Increased doses of ECU may be necessary to maintain therapeutic blood levels of ECU and full complement blockade, especially if the intestinal disease is not under control. Atypical hemolytic-uremic syndrome (aHUS) is a severe condition. Early and appropriate diagnosis and treatment are needed to avoid chronic kidney injury . PatientHere we report the case of a girl followed between the ages of 13 and 18 years with CD and aHUS. The patient was born after an uneventful twin pregnancy. Her twin brother has severe CD with complications necessitating repeated surgery for abdominal abscesses and fistulas. The first symptoms in our patient started in 2014 at the age of 13 with recurrent diarrhea, mucus in the stool, weight loss, and anemia. Initial laboratory data suggested CD  IgG were elevated ). The parents did not agree to performing an endoscopy at this stage.In parallel, the patient had anemia, normal thrombocytes, proteinuria—reaching nephrotic range—and hematuria, while serum creatinine started to increase rapidly (223 micromole/L). Abdominal ultrasound showed enlarged hyperechogenic kidneys. Kidney biopsy was performed. Two days after the kidney biopsy, the complete clinical picture of aHUS developed with thrombocytopenia, hemolytic anemia, low haptoglobin levels, acute kidney impairment (serum creatinine 263 micromole/L), and high lactate dehydrogenase levels. Indeed, kidney histology confirmed the presence of thrombotic microangiopathy.2) and low thrombocyte levels before any knowledge of the patient’s genetic background. Transfusions of fresh frozen plasma , therapeutic plasma exchange sessions (4 times), and intravenous steroid shots  were performed to eliminate eventual anti-factor H antibodies.After Enterohemorrhagic E. coli (EHEC)–induced HUS diagnosis was ruled out, complement and genetic diagnostics were performed to establish the etiology of aHUS. Treatment of TMA was urgent due to rapidly decreasing kidney function  became available and was started with an initial dose of 900 mg (for her weight of 38 kg).Complement measurements ) were performed. Total classical  and alternative pathway  activities and free serum ECU (in-house enzyme-linked immunosorbent assay from 2016 onward) levels were performed systematically directly before starting ECU infusion every 2 weeks. Fully suppressed CH50 and ALT activities were defined as an activity below 10%. Serum ECU concentrations of 50–100 microgram/mL were considered to achieve complete complement blockade [Signs of effective complement blockade were detected without delay. Classical and alternative pathway activities were depleted. Low C3 and C4 levels, as signs of severe complement consumption, normalized . Lower ADAMTS13 activity values (54–77%), as a sign of secondary endothelial damage , returneR339L). In silico prediction of variant pathogenicity was assessed, showing that the serine protease domain of human complement factor I was affected. In addition, CFH tgtgt and MCP ggaac aHUS risk haplotypes were detected. The twin brother carries the same factor I variant (R339L), the CFH tgtgt and MCP ggaac risk haplotypes. The mother bears the same CFI mutation and the MCP ggaac risk haplotype and is heterozygous for the CFH tgtgt risk haplotype. Family history was negative for aHUS despite this genetic background.A thorough genetic study of the complement system was performed and revealed the presence of a previously undescribed “likely pathogenic” factor I variant . Unexpectedly, a remarkable intra-patient variation of complement inhibition was observed. In spite of continuous complement-blocking therapy, peaks of complement pathway activities  were observed, the blockade was incomplete, and a strong association with the recurrence of bowel symptoms was found. In response, first the ECU dose was increased  assuming that the patient needed a higher dose due to weight gain. However, complement blockade was reduced even at higher doses during CD relapse. An ileum biopsy performed in April 2016 showed the typical histological picture of CD. Conventional treatment for CD  was started, with a rapid remission of CD. Non-compliance with conventional therapy resulted in CD recurrence in October 2017, coupled with the development of a perianal fistula. Biological treatment with adalimumab, a fully human, high-affinity, recombinant anti-tumor necrosis factor alpha monoclonal antibody [Figure To determine the cause of this intra-patient variability, other potential factors were analyzed. Proteinuria had not been detected since 2015, and intestinal bleeding was not significant. Free ECU serum levels were analyzed shortly before the next ECU dose from 2016 onward. Serum levels decreased to subtherapeutic concentrations (< 8–10 microgram/mL) despite regular dosing. As shown in Fig. This is a case of a pediatric patient who developed aHUS with acute kidney injury during an episode of CD, requiring long-term complement-blocking therapy. During long-term ECU treatment, a considerable intra-patient variability in the degree of complement blockade was observed along with incidents of CD. The ECU doses were adjusted for weight as recommended, but were too low for the patient with an ongoing inflammatory intestinal disease. Relapses and remissions induced characteristic fluctuations in the efficacy of complement blockade, with apparent ECU consumption during CD incidents. The temporary decrease in the effectiveness of the blockade was not associated with the return of the aHUS.ECU is a humanized monoclonal antibody against complement C5 , 4. A nuIn addition, our patient had the same genetic background as her twin brother, both patients had CD, but only the girl developed aHUS. How far the likely pathogenic CFI variant had a role in the occurrence of aHUS in our patient with CD and in the remission of aHUS under ECU is uncertain.To our knowledge, five aHUS cases associated with inflammatory bowel disease have been published in the literature –15. ImpoIn conclusion, Crohn’s disease can be one of the possible causes of secondary aHUS in children. In aHUS cases secondary to CD, close monitoring of both complement inhibition and serum ECU levels is needed as intestinal disease can interfere with complement-blocking treatment. Increased doses of ECU may be necessary to maintain therapeutic blood levels of ECU and full complement blockade, especially if the intestinal disease is not under control.


Sleep and body temperature are tightly interconnected in mammals: warming up our body helps to fall asleep and the body temperature in turn drops while falling asleep. The preoptic area of the hypothalamus (POA) serves as an essential brain region to coordinate sleep and body temperature. Understanding how these two behaviors are controlled within the POA requires the molecular identification of the involved circuits and mapping their local and brain-wide connectivity. Here, we review our current understanding of how sleep and body temperature are regulated with a focus on recently discovered sleep- and thermo-regulatory POA neurons. We further discuss unresolved key questions including the anatomical and functional overlap of sleep- and thermo-regulatory neurons, their pathways and the role of various signaling molecules. We suggest that analysis of genetically defined circuits will provide novel insights into the mechanisms underlying the coordinated regulation of sleep and body temperature in health and disease. In homeothermic animals, sleep, and body temperature are tightly interconnected. The ambient and body temperature influence the amount of sleep. Warming up the body such as taking a warm bath or passive heating promotes sleep . AnimalsThe preoptic area of the hypothalamus (POA), located in the anterior part of the hypothalamus, is known to be critical for both thermoregulation and sleep. In addition, it is also involved in regulating energy homeostasis, parenting, and sexual behaviors, each of which is controlled by dedicated circuits . HoweverThe recent development of neuroscience tools for circuit analysis has significantly helped to identify the specific cell types and regions within the POA that are crucial for sleep and body temperature regulation . We willin vivo the existence of neurons that are active during rapid eye movement (REM) and/or NREM sleep  in sleep control, a retrograde lentivirus with Cre-dependent expression of channelrhodopsin-2 (ChR2) was injected into the TMN , corticotropin-releasing hormone (CRH), and tachykinin 1 (TAC1) are highly enriched in GABAPOA→TMN neurons, and optogenetic and chemogenetic manipulation demonstrated that these subpopulations of POA neurons are indeed sleep-promoting and necessary for sleep.A recent study used cell type and projection specific targeting and gene profiling to identify molecular markers that are enriched in the POA neurons that promote sleep and are sleep-active . Such sl the TMN . Optogengulation . Optrodegulation . The firin situ hybridization (MERFISH), a recent study revealed that the POA contains a multitude of different cell populations (Using single-cell RNA sequencing (scRNA-seq) together with multiplexed error robust fluorescence ulations . This apulations . This stPreoptic area of the hypothalamus neurons project to multiple brain regions that are involved in various behaviors . In addiA receptor-mediated synaptic transmission , and monosynaptically restricted rabies tracing confirmed that they innervate hypocretin/orexin (Hcrt/Orx) neurons in the LH , 2018. Fsmission .To identify inputs to the VLPO, the retrograde tracer cholera toxin B (CTB) was injected into the VLPO and CTB-labeled neurons were examined throughout the whole brain . CTB conin vitro using slice physiology. Putative sleep neurons were identified by the presence of low-threshold spikes (LTSs)  and acetylcholine. Some of the LTS cells are either excited or inhibited by serotonin (5-HT), but not histamine (2-adrenoreceptors (The impact of monoamines on the activity of putative sleep neurons in the POA was studied s (LTSs) . Two-thiistamine , 2005. Aistamine . Thus, peceptors , 1995. Teceptors , it is ieceptors .The POA also receives major inputs from various hypothalamic regions . Histamiin vivo (Adenosine has been extensively studied as an endogenous somnogen. The adenosine level increases with prolonged wakefulness and decreases throughout sleep . During in vivo .2 (PGD2) is another powerful sleep-promoting substance. PGD2 injection into the subarachnoid space promotes NREM sleep, and injecting it just anterior and ventral to the VLPO induces its largest sleep promoting effect  of mice . This efAdcyap1, also known as PACAP) and brain-derived neurotrophic factor (Bdnf) (VMPOPACAP/BDNF) (PACAP/BDNF neurons are activated in response to warm ambient temperatures (30–42°C) or injection of capsaicin, a transient receptor potential (TRP) V1 agonist, but are non-responsive to cold temperatures, the TRP M8 agonist icilin or several non-thermal stimuli. Whole-cell patch clamp recordings from acute brain slices showed that VMPOPACAP/BDNF neurons are not more thermosensitive than neighboring cells indicating they may not be directly activated by local heat. The peripheral injection of capsaicin rapidly activated VMPOPACAP/BDNF neurons suggesting that TRPV1 sensory fibers may influence their activity. These results suggest that VMPOPACAP/BDNF neurons receive temperature signals from peripheral sensory neurons. Optogenetic stimulation of these neurons promoted hypothermia and induced thermoregulatory behaviors such as nest building. The majority of VMPOPACAP/BDNF neurons are GABAergic, and their major projection target is the DMH  . Fiber p the DMH . Taken t2)-induced fever in vivo. TRPM2 is broadly expressed throughout the POA, whereas PACAP/BDNF expression is localized within the VMPO in mice (Another study discovered the TRPM2 ion channel acts as a heat sensor and thus serves as a marker for WSNs in the POA of mice . TRPM2 i in mice . Chemoge in mice . Activat in mice .POA neurons) is involved in thermoregulation of mice (POA neurons are activated in response to warm temperatures (30°C), but not upon cold exposure (4°C). Activation of LepRbPOA neurons in mice decreased the core body temperature, locomotor activity, and energy expenditure as measured by their oxygen consumption. In addition, stimulation of LepRbPOA neurons induced postural extension, a behavioral response to high ambient temperature. The LepRbPOA neurons that are responsible for thermoregulation are mainly glutamatergic and send stimulatory signals to the DMH/dorsal hypothalamic area and rRPa in mice  has been shown to mediate energy expenditure, and a subpopulation of POA neurons expressing LepRb  into the MPA or VLPO , which control fasting-induced torpor in mice. Activating avMLPAAdcyap1 neurons decreased the body temperature and motor activity whereas inhibition disrupted torpor induction, maintenance, and arousal. Fiber photometry showed that their activity increased in response to cold exposure (10°C) and changed during the torpor-like state; their baseline level decreased, while the frequency of Ca2+-dependent events increased. Inducing a torpor-like state in non-torpid species such as human patients experiencing trauma from cardiac arrest, organ transplantation, major cardiac, and brain surgery could provide a tool to reduce organ damage (Torpor and hibernation are adaptive behaviors that allow homeothermic animals to reduce their body temperature, metabolic rate, and activity in order to survive in harsh environments with low food supply or cold temperatures . The labsumption . Two recsumption  and inje or VLPO  also ind or VLPO . Hrvatinn damage . Centraln damage , 2016.2 synthesis, suggesting that fever is mediated by PGE2 (2 into the VMPO induces fever and c-Fos expression in rats, demonstrating that the VMPO is an important site through which PGE2 promotes fever (2 has been shown to act on warm-sensitive thermoregulatory neurons in the hypothalamus (2 activates one of its receptors, EP3R, expressed in excitatory VMPO/MnPO neurons to mediate the fever response in mice (2 action through the EP3R receptors in VMPO/MnPO glutamatergic neurons and is eliminated when these neurons are ablated. Glutamatergic EP3R-expressing POA neurons directly project to the rRPa and this pathway may mediate inflammatory fever (Fever, a significant body temperature increase in response to infections and inflammation is an evolutionarily conserved phenomenon and necessary to help the immune system fight infections and inhibit the growth of microorganisms . During  by PGE2 . Microines fever . In partthalamus . PGE2 ac in mice . LPS-indry fever . Taken tThe POA comprises many subdivisions, and the recent identification of novel molecular markers that specifically label neurons regulating sleep or body temperature helped to precisely define the subregions and cell types controlling these fundamental behaviors. Furthermore, newly developed tools greatly improved the temporal and spatial resolution for recording and manipulating these neurons providing important insights into the underlying circuit mechanisms .However, the extent of anatomical overlap between warm-sensitive and sleep-promoting POA neurons is still largely unknown. Consequently, the projection pattern of such neurons and their interaction with other regions involved in sleep and temperature control is still unknown. An interesting hypothesis to test in future studies is whether neurons involved in both behaviors send axonal projections to both sleep and temperature-regulatory downstream areas. The location of newly identified sleep and thermoregulatory neurons partially overlap. Previous studies indicated that a subset of POA neurons activated by local warming becomes activated during NREM sleep . It is uPreoptic area of the hypothalamus sleep and thermoregulatory neurons express a variety of peptidergic markers as well as receptors and channels . An impoClock develop obesity and metabolic syndromes suggesting that this circadian clock gene plays an important role in energy balance (In addition to thermoregulation, sleep is also closely interconnected with metabolism. During NREM sleep, the energy metabolism is strongly reduced. Chronic sleep fragmentation in mice prevents the reduction of the brain temperature during sleep and leads to increased food intake and glucose intolerance suggesting that poor-quality sleep may contribute to development of metabolic dysfunctions . Similar balance . For fut balance .Different types of sleep disorders, especially those associated with aging, may be influenced by altered body temperature rhythms or dysregulation of vasodilation resulting in an impairment to prepare the body for sleep . DelayedBoth authors wrote the manuscript, made the figures, 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.


According to international guidelines, recurrent inguinal hernia should be treated by a surgical approach opposing of the primary strategy (anterior–posterior or posterior–anterior). However, recent evidence demonstrates feasibility and safety of re-laparoscopic repair of recurrent inguinal hernia after primary laparoscopy. For such a strategy, correct identification of anatomical structures is challenging, but absolutely crucial for a satisfactory postoperative result. This case of an unrecognized sliding hernia of the sigmoid colon during re-laparoscopy highlights that a precise physical examination as well as an extended preoperative radiological workup  should be considered prior to re-laparoscopy of recurrent inguinal hernia. Nearly 20 million primary inguinal hernia operations are performed globally and annually. A recurrence rates of up to 12% have been reported with <50% being reoperated. Diagnosis of a recurrent hernia can be difficult, especially when clinical symptoms are equivocal. Physical examination combined with sonography represents the standard approach. In unclear cases, magnetic resonance imaging (MRI) or computed tomography (CT) may be performed additionally [The European Hernia Society and HerniaSurge Group guidelines recommend a change of the surgical strategy for reoperations, i.e. the use of posterior repair in case of primary anterior repair and vice versa , 2. HoweHerein, we demonstrate potential pitfalls of re-laparoscopic repair of recurrent inguinal hernia and highlight advantages of an extended preoperative radiological workup.A male patient (51 years) underwent bilateral transabdominal preperitoneal (TAPP) repair for a large left-sided sliding hernia (sigmoid colon) and a small right-sided medial inguinal hernia 7 months ago. A large symptomatic recurrence on the left side was clinically confirmed.Considering the patients’ comorbidities , a re-TAPP was performed to avoid possible wound complications. The recurrent hernia was presumably confirmed as a lateral inguinal hernia. After dissecting dense fibrotic scar tissue around the previous mesh preperitoneally, the hernia sac was released. A large spermatic cord lipoma was found and dissected  and 2. AHowever, the patient experienced persistent swelling in the left groin. To confirm the early re-recurrence hernia, CT of the abdomen revealed a left sliding hernia with sigmoid colon .The patient agreed to proceed with diagnostic laparoscopy for a re-re-repair of the left inguinal hernia. Intraoperatively, a loop of the sigmoid colon, which previously was hidden by colonic adhesions, was fixed into a hernia orifice lateral–caudal of the preperitoneal mesh . After rThe perioperative course was uneventful with discharge at day 3. No recurrence was seen 6 months postoperatively.Surgery of a symptomatic inguinal hernia is one of the most often performed procedures worldwide. The primary laparo-endoscopic approach ) is considered the standard procedure by many and has replaced the primary anterior technique with or without mesh  or Shouldice). Although laparo-endoscopic techniques have decreased the recurrence rates significantly , rates oBefore reoperation of a recurrent inguinal hernia, physical examination and an ultrasound of the inguinal region are recommended. In addition, MRI or CT may be added in unclear cases.There is a paucity of studies investigating the extent of preoperative radiological imaging in cases of recurrent inguinal hernia and data to support an extensive workup are scarce. A recent retrospective analysis showed 97% positive prediction of diagnosing recurrent inguinal hernia by physical examination only, suggesting that an additional ultrasound may not always be necessary . As the The literature to support diagnosis of primary and recurrent inguinal hernia remains equivocal. Low-dose CT of the inguinal region has gained importance. Advantages include evaluation of hernia content, simultaneous investigation of the contralateral side and differentiation of other causes of inguinal swelling. In this case, a preoperative CT would have shown the sliding hernia with sigmoid colon and a sufficient surgical dissection during the first re-laparoscopy may have been performed, omitting re-re-laparoscopy.Surgery for recurrent inguinal hernia can be challenging. An open anterior approach for the treatment of a recurrent inguinal hernia originally repaired by a laparoscopic technique, and vice versa, is recommended by international guidelines . NonetheOur patient suffered from obesity World Health Organization grade II in combination with metabolic comorbidities. Thus, it was reasonable to expect him to benefit from minimal invasive approach. After the dissection of the scar tissue, a large spermatic cord lipoma was detected and mistakenly identified as cause of hernia recurrence. No further dissection of the preperitoneal space was undertaken. This is not in accordance with recently published literature advising a dissection of the preperitoneal layer to at least 2 cm below the pubic symphysis . This reThe re-re-laparoscopic operation confirmed a sliding hernia of the sigmoid. A complete dissection of the preperitoneal tissue was necessary. A new mesh covering the indirect, direct and femoral triangle with overlapping of the pubic symphysis was placed.Alternative may have been a laparoscopic closure of the hernia gap, insertion of a mesh patch and preservation of the former mesh as recently proposed .Although, there is no data available, robotic approach in inguinal hernia surgery may represent a beneficial alternative as it is characterized by enhanced visibility, sole control by surgeon and improved dexterity, which may refine tissue preparation even in technically demanding situations .In summary, we present a case of a missed sliding hernia of the sigmoid during re-laparoscopy of a recurrent inguinal hernia following primary TAPP. We recommend extended radiological imaging including CT or MRI prior to revisional surgery. Re-laparoscopy is feasible and associated with all benefits of minimal invasive surgery. A thorough tissue preparation needs to be applied to recognize important anatomical landmarks. Robotic inguinal hernia surgery may have an important role in revisional surgery.None declared.


Previous research establishes that hurricanes adversely affect nursing home (NH) resident health but specific causal pathways are still unclear. We combined power outage data with Medicare claims to determine the effects of power loss from Hurricane Irma(2017) among NH residents in Florida. Out of 580 facilities, 289 reported power loss. These facilities had higher star ratings; higher beds counts, and were preferentially in the Southeast region of Florida compared to facilities without outages. There were 27,767 residents living in a NH without power. They were comparable in characteristics to residents that did not lose power . We ran adjusted generalized linear models with robust standard errors, clustering for NH. We found power loss was associated with a trend towards increased odds of mortality within 7-days  and 30-days  post-storm, but not with hospitalization. Future research should investigate the time-specific effects of power outages.


Antibody-mediated disorders of the central nervous system (CNS) are increasingly recognized as neurologic disorders that can be severe and even life-threatening but with the potential for reversibility with appropriate treatment. The expanding spectrum of newly identified autoantibodies targeting glial or neuronal  antigens and associated clinical syndromes  has increased diagnostic precision, and allowed critical reinterpretation of non-specific neurological syndromes historically associated with systemic disorders . The intracellular vs. cell-surface or synaptic location of the different neural autoantibody targets often helps to predict the clinical characteristics, potential cancer association, and treatment response of the associated syndromes. In particular, autoantibodies targeting intracellular antigens  are often associated with cancers, rarely respond well to immunosuppression and have a poor outcome, although exceptions exist. Detection of neural autoantibodies with accurate laboratory assays in patients with compatible clinical-MRI phenotypes allows a definite diagnosis of antibody-mediated CNS disorders, with important therapeutic and prognostic implications. Antibody-mediated CNS disorders are rare, and reliable autoantibody identification is highly dependent on the technique used for detection and pre-test probability. As a consequence, indiscriminate neural autoantibody testing among patients with more common neurologic disorders  will necessarily increase the risk of false positivity, so that recognition of high-risk clinical-MRI phenotypes is crucial. A number of emerging clinical settings have recently been recognized to favor development of CNS autoimmunity. These include antibody-mediated CNS disorders following herpes simplex virus encephalitis or occurring in a post-transplant setting, and neurological autoimmunity triggered by TNFα inhibitors or immune checkpoint inhibitors for cancer treatment. Awareness of the range of clinical and radiological manifestations associated with different neural autoantibodies, and the specific settings where autoimmune CNS disorders may occur is crucial to allow rapid diagnosis and early initiation of treatment. The spectrum of immune-mediated neurologic disorders is rapidly expanding with the growing understanding of diverse disease mechanisms that can lead to central and/or peripheral nervous system inflammation , 2. AntiIn this article, we will review the main clinical syndromes and MRI characteristics associated with neural autoantibodies directed against CNS antigens, and discuss commonly encountered challenges for diagnosis and management. Since not all target antigens are exclusively expressed in the CNS, certain neural autoantibodies are associated with complex clinical syndromes characterized by concomitant CNS and extra-CNS manifestations. On the contrary, neurological syndromes associated with autoantibodies exclusively targeting neuromuscular structures  will not be discussed in this article; yet share many similarities in terms of pathophysiology and management.Neural autoantibodies are generally classified on the basis of the cellular location of their target antigens, intracellular vs. cell-surface or synaptic . As a geThe cellular location of the target antigens also has clinical implications. Antibodies targeting intracellular antigens are rare in children, have a stronger cancer association , and generally respond less well to immunosuppressive treatments ; while aIdentification of specific neural autoantibodies in patients with suspected autoimmune neurologic disorders provides important advantages to the clinician, including:1) Diagnostic confirmation—confirming a diagnosis of autoimmune neurologic disorder is crucial to avoid treatment delays, inappropriate treatments for other disorders, and unnecessary diagnostic procedures  ( Diagnost(2) Treatment precision—specific treatments often differ between different antibody-mediated disorders, but also from other immune-mediated disorders without specific neural autoantibodies. For instance, disease modifying agents commonly used to treat multiple sclerosis  may be ineffective or even worsen AQP4 and MOG antibody-associated disorders –22;(3) Cancer prediction—certain neural autoantibodies are strongly associated with specific types of cancer, and can appear months to years before the cancer becomes clinically manifest allowing early diagnosis and treatment .However, neural autoantibody testing is not flawless, and its reliability depends on a number of factors including the type of laboratory assay used for detection, the characteristics of the population tested, and the clinical setting.There are two major requirements for a correct diagnosis of antibody-mediated CNS disorder to be made: (1) a reliable identification of one or more specific neural autoantibodies; and (2) a compatible clinical-MRI phenotype.3 with or without oligoclonal bands is useful to help distinguish from non-inflammatory mimics when present, but its frequency might decrease in the elderly or depend on the antibody associated  and indistinguishable from those of other non-inflammatory etiologies  so that exclusion of alternative and more common diagnoses is mandatory –25. An asociated . The detsociated . Examplesociated .Limbic encephalitis is one of the most common phenotypes and can be seen in association with a wide variety of neural autoantibodies ..Limbic etibodies . Brain Mtibodies , with ortibodies , neoplastibodies , and seitibodies . AutoimmProgressive encephalopathy with rigidity and myoclonus (PERM) is a rare phenotype characterized by the occurrence of muscular rigidity, spasms, and myoclonus in association with encephalopathy and usually accompanied by a normal MRI head and spine , insomnia, hyperhidrosis and is frequently seen with LGI1 and Caspr2 autoantibodies  are short-lasting dystonic movements primarily affecting the arm and face on one side, with a high daily frequency (up to 40–60 times per day) (per day) . These sper day)  or T2-weper day) .Opsoclonus-myoclonus syndrome is characterized by rapid, multi-directional eye movement and diffuse muscle jerks, often in association with ataxia. This syndrome can be seen in children with neuroblastoma and ANNA1/anti-Hu antibodies but also in adults with breast or lung cancers and ANNA2/anti-Ri antibodies  or ADEM-like phenotypes are characterized clinically by encephalitis and/or myelitis accompanied by large T2-hyperintense demyelinating lesions in multiple CNS regions on MRI . Large T2 hyperintensities with indistinct margins can be encountered and involve the middle cerebellar peduncle , 64OpticMyelitis lesions that are longitudinally-extensive  are also typical of both AQP4 antibodies  is a less common, but quite characteristic clinical-MRI phenotype seen with MOG autoantibodies  is found in up to one third of patients with NMDAR autoantibodies , althoug18F-fluorodeoxyglucose positron emission tomography (FDG-PET) is more sensitive than brain MRI in patients with autoimmune encephalitis and can show abnormal hypometabolism, hypermetabolism, or both in a diffuse fashion or restricted to certain brain areas  . Brain oe.g., mammogram, gynecologic/scrotal ultrasound) should not be overlooked. Certain autoantibodies have a strong association with specific types of cancer  used for testing, and the setting in which the test is performed . DiffereHuman embryonic kidney (HEK) cells are transfected with genes encoding the target antigen of interest, which is expressed on the surface of the cell membrane. This technique allows binding of the human autoantibodies to their target antigen in its native conformational form and is generally the assay of choice for antibodies targeting cell surface/synaptic antigens  92, 93), 9392, 9These techniques allow recognition of solubilized antigens by their specific human antibodies but not in a native conformational form and are commonly used to detect antibodies targeting intracellular antigens , 97. TheLive neuronal cell cultures can also be used to confirm autoantibody binding to surface antigens .Unfortunately, TIF is routinely available in only few specialized laboratories worldwide and thus many laboratories utilize single methods for detection. Novel cell-based assays are now available which have much improved sensitivity and specificity and have a low risk of false positive results (0.2%) when compared to older generation techniques in which false positives are much more common (up to 6%) .When possible, neural autoantibodies should be tested in both serum and CSF to maximize sensitivity, although certain autoantibodies are preferentially detected in either one of these biospecimens . NMDAR aDespite a high accuracy reported for most of the commercially available assays, a positive autoantibody result does not necessarily imply that the patient has an autoimmune neurologic disorder. As with any diagnostic biomarker, sensitivity, and specificity are intrinsic characteristics of the test and measure the quality of the assay and the testing laboratory, but are less reflective of the setting in which the test is performed. A more useful parameter in clinical practice is the positive predictive value (PPV), which is the ratio between true positives and total positive results obtained by testing a given population, and is highly dependent on the frequency of the disease of interest in the population tested and the test ordering practices . This isHigh autoantibody titers are less likely to be seen with false positive results, but they still do not guarantee 100% specificity. GAD65 autoantibodies are a typical example since they can be non-specifically found at low titers in diabetic patients or patients without neurologic deficits. However, the detection of high-titer GAD65 autoantibodies in the serum and/or CSF of neurologic patients is not sufficient for a diagnosis of GAD65 autoimmunity in the absence of one of the compatible clinical phenotypes of stiff-person spectrum, autoimmune ataxia or autoimmune epilepsy. In a recent study of 323 patients with high-titer GAD65 autoantibodies seen at Mayo Clinic, one third had an alternative diagnosis . AwareneSpecific diagnostic criteria have been published to guide clinicians during the diagnostic process in patients with suspected autoimmune encephalitis . Based oAmong autoimmune demyelinating CNS disorders, international consensus diagnostic criteria exist for neuromyelitis optica spectrum disorders (NMOSD) with AQP4 autoantibodies . AlthougOtherwise unexplained neurological manifestations in patients with systemic autoimmune/inflammatory disorders have frequently been regarded as being part of the systemic syndrome, especially in the past. Although neurological manifestations of systemic disorders are possible  , the conSpecific clinical settings are increasingly recognized to be at higher risk for development of autoimmune CNS disorders. These include:HSV encephalitis often present with symptoms/signs of mesial temporal lobe involvement and can be difficult to differentiate from autoimmune limbic encephalitis. However, autoimmune encephalitis may develop after HSV encephalitis  in up to 27% of patients. These forms are generally associated with neural autoantibodies (frequently targeting NMDAR) and should be suspected in patients with HSV encephalitis who show clinical and/or MRI worsening after an initial improvement with anti-viral therapy .Autoimmune CNS disorders may rarely paradoxically arise in post-transplant patients despite ongoing immunosuppressant medication use to prevent organ rejection. It is possibly due to imbalance between B- and T-cell as anti-rejection immunosuppression are predominantly T-cell targeting treatments. Testing neural antibodies in this scenario is reasonable and the addition of antibody depleting treatments  in addition to maintenance anti-rejection immunosuppression should be considered .TNFα inhibitors are used for treatment of many systemic  and neurologic  chronic inflammatory disorders. CNS inflammatory disorders of any type, with or without accompanying neural autoantibodies, may develop in patients treated with these drugs  with a higher frequency compared to untreated patients .ICI act by enhancing the immune response against cancers and are increasingly approved for treatment of different tumors. A common and unwanted effect of these drugs is autoimmunity, potentially directed against any organ –125. CNSCases of neurological autoimmunity including antibody-associated encephalitis has been reported in multiple sclerosis patients treated with the CD52  or CD25  protein inhibitors –129.Diverse neurologic manifestations have been described in patients with SARS-CoV-2 infection during the COVID-19 pandemic, including antibody-associated CNS disorders , 131. AlCurrent treatment strategies for antibody-mediated neurologic disorders are mostly based on anecdotal experience, expert opinion and data from retrospective case series. The treatment approaches differ based on the specific autoantibody detected, associated syndromes, and phase of the disease (acute/active CNS inflammation vs. remission). The rarity of these disorders has hampered the development of randomized controlled clinical trials in autoimmune neurologic disorders. However, the recent successful randomized controlled clinical trials of a number of attack-prevention targeted treatments in AQP4-IgG seropositive NMOSD has provided a platform for future clinical trials in antibody mediated syndromes. However, for most autoimmune neurologic disorders there are no proven treatments but some general principles have been developed that can guide treatment decisions when definitive scientific evidence is lacking.The treatment approach for acute exacerbations of antibody-mediated CNS disorders is similar for the different underlying autoantibodies and includes one or more of:Intravenous methylprednisolone . The equivalent dose of 1,250 mg of prednisone once daily for 5 days can be considered as an alternative and has been utilized in some patients when there have been difficulties in arranging intravenous infusions . Due to the lower risk of potential adverse effects, IVMP is generally preferred and often sufficient to improve symptoms as a monotherapy. Common adverse events related to IVMP include hyperglycemia in patients with diabetes or impaired glucose tolerance, and corticosteroid-induced psychosis.Intravenous immunoglobulins (IVIg–0.4 g/Kg once daily for 5 days).Plasma exchange (PLEX–1 exchange every other day for 5–7 exchanges).IVIg and PLEX are generally used as add-on to IVMP for more severe cases. IVIg may be contraindicated in patients with kidney disease, IgA deficiency, and is associated with an increased risk of thrombosis while PLEX may induce hypotension and should be avoided in patients with low baseline blood pressure and is complicated by the frequent need for central line placement. PLEX has been shown to be highly effective as acute monotherapy in patients with severely disabling idiopathic inflammatory diseases of the CNS with a randomized, sham-controlled, double-blinded trial. The study was published in 1999 and is likely that many of included patients with severe attacks of demyelination considered idiopathic at the time now would test positive for AQP4 or MOG autoantibodies . More reOne small randomized placebo-controlled trial of IVIg in 17 patients with autoimmune epilepsy associated with LGI or Caspr2 autoantibodies showed a significant difference in seizure reduction compared to placebo . In our A multicenter observational study analyzed treatment responses and outcomes in 501 patients with NMDAR autoantibody-positive encephalitis for 2 years . ImproveTreatment of CNS autoimmunity secondary to TNFα inhibitors or ICI should ideally require a combination of traditional acute immunotherapies  and withdrawal of the triggering agent. In cancer patients treated with ICI, however, the risk and benefits of ICI withdrawal should be carefully weighted based on cancer status and the severity of neurologic manifestations . ImproveLong-term immunosuppression aims to prevent relapses after the acute attacks, or worsening of neurological manifestations in patients with a chronic progressive course associated with certain autoantibodies. The choice of the immunosuppressive agent is mostly based on the type of autoantibody detected and safety profile of the drug. Neurological CNS syndrome associated with autoantibodies targeting cell-surface proteins are generally treated with antibody-depleting or B-cell-depleting agents  while syndromes with autoantibodies targeting intracellular antigens that are thought to be part of a predominantly cytotoxic process are preferentially treated with agents depleting all types of immune cells  although some overlap exists in treatment regimens. However, a reduced effectiveness of B-cell depleting agents with autoantibodies targeting intracellular antigens has not been proven and the mechanism of action of rituximab might also indirectly involve T-cells and depletion of B cells may also reduce T cell function and depleting T-cells may reduce B-cell function. Commonly used long-term immunosuppressants and related administration regimens are listed below:Rituximab is an anti-CD20 monoclonal antibody. It is widely used worldwide due to the good tolerability, limited risk of adverse reactions, and fast onset to action. There are two main regimens of intravenous administration for rituximab in adults: (1) Induction with two 1 g-infusions 2 weeks apart and the same regimen every 6 months; and (2) induction with 375 mg/m2 of body surface area/week for 4 consecutive weeks, then periodic reinfusions with half of the initial dose (1 reinfusion/week for two consecutive weeks). The timing of reinfusions can be either fixed , or guided by monthly/bimonthly monitoring of CD19-positive cell count (in this case the drug is reinfused when the proportion of CD19 positive cells increases >1%). Side effects include infusion reactions and risk of infections sometimes in the setting of a secondary hypogammaglobulinemia; thus, monitoring of total IgG levels 6 monthly can be considered.Azathioprine is an antagonist of purine synthesis and consequently DNA/RNA production for the proliferation of white blood cells. It is generally administered orally with a recommended total daily dose of 2–3 mg/Kg. Azathioprine commonly takes 6–8 months to become effective so that a prolonged, concomitant taper of oral steroids is often needed in conjunction. Side effects include rash, infection, macrocytic anemia, hypersensitivity reactions, pancreatitis, elevated liver enzymes and increased risk of tumors . Testing thiopurine methyltransferase (TMPT) enzyme activity before starting is mandatory and those with deficiency are at increased risk of severe bone marrow toxicity and in this population the medication should be avoided.Mycophenolate Mofetil is another oral inhibitor of purine metabolism, mainly acting on lymphocytes. The recommended starting dose in adults is 500 mg twice daily and increasing to a goal dose of 1,000 mg twice daily. This drug becomes effective in ~2–3 months. Side effects include GI disturbance, infections, and increased risk of tumors .Cyclophosphamide acts by inducing cell apoptosis via induction of irreversible DNA alterations. It is generally reserved for severe cases refractory to other immunotherapies due to the strong immunosuppressive effect and greater risk of adverse events which include nausea and vomiting, alopecia, hemorrhagic cystitis, agranulocytosis, infertility and increased risk of tumors . The recommended dose in adults is 1,000 mg/m2 monthly intravenously or 1–2 mg/kg once daily by mouth. In general, treatment is limited to 6 months after which transition can be considered to one of the other immunosuppressants mentioned above.Novel therapies targeting key specific proteins of the inflammatory cascade have also been proposed as promising and highly effective treatments for antibody-mediated CNS disorders and can be considered for refractory cases. These include inhibitors of certain interleukins or their receptors , and specific complement proteins  , 140. HeA number of monoclonal antibodies have recently been proven to be highly effective for relapse prevention in patients with AQP4 autoantibodies by randomized clinical trials, including eculizumab (anti-C5 complement protein) , satraliIn patients with paraneoplastic antibody-mediated CNS disorders, cancer removal should be prioritized even in patients with severe acute disability. Early cancer removal has been associated with improved outcomes and better treatment response . Other cAR  , 150. DeAR . ManagemAmong autoantibodies associated with CNS demyelination, those targeting AQP4 are typically associated with a worse outcome given the destructive nature of their clinical attacks which leads to a stepwise disability accumulation , 157. ClThe outcome is often favorable also in patients treated with ICI who develop autoimmune CNS disorders, although severe residual disability can be observed in approximately one third of patients . Older aIn conclusion, antibody-mediated CNS disorders are heterogeneous with variable clinical-MRI characteristics and prognoses based on the specific underlying autoantibody. Identification of one or more neural specific autoantibodies in the serum and/or CSF of patients with a compatible clinical phenotype confirms the diagnosis but false positive results may occur especially with indiscriminate testing, which could lead to inappropriate treatment. General treatment principles exist for antibody-mediated CNS syndromes exist but most lack evidence to support their use and are often broad immunosuppressants rather than targeted treatments. Future directions for this field include developing more targeted treatments for each antibody-mediated syndrome using the success in aquaporin-4 positive NMOSD as a guide on how specific targeted treatments can be developed, proven to work and made available for patient care.This manuscript was entirely planned and drafted by ES and EF, including drafting of the Figures and Tables. Both 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.