Traumatic brain injury (TBI) is a leading global cause of death and disability. Here we demonstrate in an experimental mouse model of TBI that mild forms of brain trauma cause severe deficits in ...meningeal lymphatic drainage that begin within hours and last out to at least one month post-injury. To investigate a mechanism underlying impaired lymphatic function in TBI, we examined how increased intracranial pressure (ICP) influences the meningeal lymphatics. We demonstrate that increased ICP can contribute to meningeal lymphatic dysfunction. Moreover, we show that pre-existing lymphatic dysfunction before TBI leads to increased neuroinflammation and negative cognitive outcomes. Finally, we report that rejuvenation of meningeal lymphatic drainage function in aged mice can ameliorate TBI-induced gliosis. These findings provide insights into both the causes and consequences of meningeal lymphatic dysfunction in TBI and suggest that therapeutics targeting the meningeal lymphatic system may offer strategies to treat TBI.
Nascent RNA profiling is growing in popularity; however, there is no standard analysis pipeline to uniformly process the data and assess quality. Here, we introduce PEPPRO, a comprehensive, scalable ...workflow for GRO-seq, PRO-seq, and ChRO-seq data. PEPPRO produces uniformly processed output files for downstream analysis and assesses adapter abundance, RNA integrity, library complexity, nascent RNA purity, and run-on efficiency. PEPPRO is restartable and fault-tolerant, records copious logs, and provides a web-based project report. PEPPRO can be run locally or using a cluster, providing a portable first step for genomic nascent RNA analysis.
Sequence‐specific transcription factors (TFs) bind DNA, modulate chromatin structure, regulate gene expression, and orchestrate transcription cascades. Activation and repression of TFs drive tightly ...controlled regulatory programs that lead to cellular processes such as differentiation. We measured chromatin accessibility and nascent transcription at seven time points over the first four hours of induced adipogenesis of preadipocytes to construct dynamic gene regulatory networks. Regulatory networks describe successive waves of TF binding and dissociation followed by direct regulation of proximal genes. We identified 14 families of TFs that coordinate with and antagonize each other to regulate early adipogenesis. We developed a compartment model to quantify individual TF contributions to RNA polymerase initiation and pause release rates. Network analysis showed that the glucocorticoid receptor and AP1 drive immediate gene activation, including induction of Twist2. Twist2 is a highly interconnected node within the network and its expression leads to repression of target genes. Although Twist2's role in adipogenesis has not been previously appreciated, both Twist2 knockout mice and Setleis syndrome (Twist2‐/‐) patients lack subcutaneous and brown adipose tissue. We found that kinetic networks integrating chromatin structure and nascent transcription dynamics identify key genes, TF functions, and coordinate interactions within regulatory cascades.
Emerging evidence suggests that the meningeal compartment plays instrumental roles in various neurological disorders, however, we still lack fundamental knowledge about meningeal biology. Here, we ...utilized high-throughput RNA sequencing (RNA-seq) techniques to investigate the transcriptional response of the meninges to traumatic brain injury (TBI) and aging in the sub-acute and chronic time frames. Using single-cell RNA sequencing (scRNA-seq), we first explored how mild TBI affects the cellular and transcriptional landscape in the meninges in young mice at one-week post-injury. Then, using bulk RNA-seq, we assessed the differential long-term outcomes between young and aged mice following TBI. In our scRNA-seq studies, we highlight injury-related changes in differential gene expression seen in major meningeal cell populations including macrophages, fibroblasts, and adaptive immune cells. We found that TBI leads to an upregulation of type I interferon (IFN) signature genes in macrophages and a controlled upregulation of inflammatory-related genes in the fibroblast and adaptive immune cell populations. For reasons that remain poorly understood, even mild injuries in the elderly can lead to cognitive decline and devastating neuropathology. To better understand the differential outcomes between the young and the elderly following brain injury, we performed bulk RNA-seq on young and aged meninges 1.5 months after TBI. Notably, we found that aging alone induced upregulation of meningeal genes involved in antibody production by B cells and type I IFN signaling. Following injury, the meningeal transcriptome had largely returned to its pre-injury signature in young mice. In stark contrast, aged TBI mice still exhibited upregulation of immune-related genes and downregulation of genes involved in extracellular matrix remodeling. Overall, these findings illustrate the dynamic transcriptional response of the meninges to mild head trauma in youth and aging.
Adipocytes contribute to metabolic disorders such as obesity, diabetes, and atherosclerosis. Prior characterizations of the transcriptional network driving adipogenesis have overlooked transiently ...acting transcription factors (TFs), genes, and regulatory elements that are essential for proper differentiation. Moreover, traditional gene regulatory networks provide neither mechanistic details about individual regulatory element-gene relationships nor temporal information needed to define a regulatory hierarchy that prioritizes key regulatory factors. To address these shortcomings, we integrate kinetic chromatin accessibility (ATAC-seq) and nascent transcription (PRO-seq) data to generate temporally resolved networks that describe TF binding events and resultant effects on target gene expression. Our data indicate which TF families cooperate with and antagonize each other to regulate adipogenesis. Compartment modeling of RNA polymerase density quantifies how individual TFs mechanistically contribute to distinct steps in transcription. The glucocorticoid receptor activates transcription by inducing RNA polymerase pause release, whereas SP and AP-1 factors affect RNA polymerase initiation. We identify
as a previously unappreciated effector of adipocyte differentiation. We find that TWIST2 acts as a negative regulator of 3T3-L1 and primary preadipocyte differentiation. We confirm that
knockout mice have compromised lipid storage within subcutaneous and brown adipose tissue. Previous phenotyping of
knockout mice and Setleis syndrome
patients noted deficiencies in subcutaneous adipose tissue. This network inference framework is a powerful and general approach for interpreting complex biological phenomena and can be applied to a wide range of cellular processes.
Guillain‐Barré syndrome (GBS) is the commonest post‐infectious polyradiculopathy. Although genetic background of the host seems to play an important role in the susceptibility to GBS, genes ...conferring major risk are not yet known. Dysregulation of Toll‐like receptor (TLR) molecules exacerbates immune‐inflammatory responses and the genetic variations within TLR pathway‐related genes contribute to differential risk to infection. The aim of this study was to delineate the impact of genetic variations within TLR2, TLR3, and TLR4 genes as well as TLR signaling pathway‐related genes such as MyD88, TRIF, TRAF3, TRAF6, IRF3, NFκβ1, and IκBα on risk of developing GBS. Fourteen polymorphisms located within TLR2 (rs3804099, rs111200466), TLR3 (rs3775290, rs3775291), TLR4 (rs1927911, rs11536891), MyD88 (rs7744, rs4988453), TRIF (rs8120), TRAF3 (rs12147254), TRAF6 (rs4755453), IRF3 (rs2304204), NFκβ1 (rs28362491), and IκBα (rs696) genes were genotyped in 150 GBS patients and 150 healthy subjects either by PCR‐RFLP or TaqMan Allelic Discrimination Assay. Genotypes of two polymorphic variants, Del/Del of rs111200466 insertion and deletion (INDEL) polymorphism of TLR2 gene and TT of rs3775290 single nucleotide polymorphism (SNP) of TLR3 gene had significantly higher frequencies among GBS patients, while the frequencies of TT genotype of rs3804099 SNP of TLR2 gene and TT genotype of rs11536891 SNP of TLR4 gene were significantly higher in controls. Gene‐gene interaction study by Multifactor Dimensionality Reduction analysis also suggested a significant combined effect of TLR2, and NFκβ1 genes on the risk of GBS. The SNPs in the IκBα and IRF3 genes correlated with severity of GBS. The genes encoding TLRs and TLR signaling pathway‐related molecules could serve as crucial genetic markers of susceptibility and severity of GBS.
The clinical success of combined androgen deprivation therapy (ADT) and radiotherapy (RT) in prostate cancer created interest in understanding the mechanistic links between androgen receptor (AR) ...signaling and the DNA damage response (DDR). Convergent data have led to a model where AR both regulates, and is regulated by, the DDR. Integral to this model is that the AR regulates the transcription of DDR genes both at a steady state and in response to ionizing radiation (IR). In this study, we sought to determine which immediate transcriptional changes are induced by IR in an AR-dependent manner. Using PRO-seq to quantify changes in nascent RNA transcription in response to IR, the AR antagonist enzalutamide, or the combination of the two, we find that enzalutamide treatment significantly decreased expression of canonical AR target genes but had no effect on DDR gene sets in prostate cancer cells. Surprisingly, we also found that the AR is not a primary regulator of DDR genes either in response to IR or at a steady state in asynchronously growing prostate cancer cells.
Our data indicate that the clinical benefit of combining ADT with RT is not due to direct AR regulation of DDR gene transcription, and that the field needs to consider alternative mechanisms for this clinical benefit.
Guillain‐Barré syndrome (GBS) is the commonest post‐infectious inflammatory peripheral neuropathy with undiscerned aetiology. The commonly reported antecedent infections implicated in India include ...Campylobacter jejuni, chikungunya, dengue, and Japanese encephalitis (JE). In this study from south India, we investigated the role of these four agents in triggering GBS. This case‐control study was performed on 150 treatment‐naive patients with GBS and 150 age and sex‐matched controls from the same community. IgM immunoreactivity for C. jejuni, chikungunya, and dengue was detected by enzyme‐linked immunosorbent assay (ELISA) in serum of patients with GBS and control subjects. Immunoreactivity against JE was detected in serum as well as cerebrospinal fluid (CSF) from patients (n = 150) and orthopaedic control (n = 45) subjects. The immunoreactivity against infections was compared between demyelinating and axonal subtypes of GBS. Overall, 119/150 patients with GBS had serological evidence of antecedent infection. Amongst those with evidence of antecedent infection, 24 (16%), 8 (5%), and 9 (6%) patients were exclusively immunoreactive to chikungunya, JE, and C. jejuni, respectively. In the remaining patients (78/119), immunoreactivity to multiple pathogens was noted. Immunoreactivity to C. jejuni infection was found in 32% of GBS patients compared to 2.7% controls (P < .001), whereas to chikungunya virus was reported in 66.7% of patients with GBS compared to 44.7% controls (P = .006). Anti‐dengue immunoreactivity was significantly associated with the demyelinating subtype of GBS. Patients positive for JE IgM (CSF) manifested demyelinating electrophysiology. In this large case‐control study, immunoreactivity against multiple infectious agents was observed in a subset of patients. Chikungunya was the commonest antecedent infection, followed by C. jejuni.
Background
Antibodies against ganglioside complexes (GSCs) are associated with various clinical features and subtypes of Guillain‐Barré syndrome (GBS).
Methods
One‐hundred patients were evaluated for ...antibodies to GSCs formed by combination of GM1, GM2, GD1a, GD1b, GT1b, and GQ1b using manual enzyme linked immuno‐sorbent assay (ELISA).
Results
Twenty‐six patients were GSC antibody‐positive, most frequent being against GM1‐containing GSC (76.9%). Gender distribution, mean age, symptom‐duration, antecedent events, electrophysiological subtypes, requirement for mechanical ventilation, and median duration of hospital stay were comparable between the GSC antibody‐positive and negative groups. There was no association between specific GSC antibody and electrophysiological subtypes or clinical variants. After controlling for false discovery rate (FDR) using the Benjamini‐Hochberg method, the number of subjects who improved in overall disability sum score, modified Erasmus GBS outcome score, and neuropathy symptom score at discharge was significantly higher in the GSC antibody‐positive group. Improvements in Medical Research Council sum scores and Hughes Disability Scale during the hospital stay between the GSC antibody‐positive and negative groups were not significantly different after controlling for FDR.
Conclusions
The GSC antibody‐positive group had better outcome at hospital discharge in some of the disability scores. Pathophysiological pathways among patients without GSC antibodies may be different and this requires further evaluation.
To analyse the pattern of cardiovascular diseases (CVDs) in COVID-19 patients admitted to tertiary cardiac care centre.
We retrospectively analysed 511 adult patients admitted between July 1, 2020, ...and November 30, 2020, with COVID-19 infection and having either new onset or pre-existing CVDs. Clinical features, electrocardiogram (ECG), echocardiography, chest X-ray, biomarkers, haematological and biochemical parameters were analysed.
The mean age of the patients was 56.62 ± 14.74 years. Male: Female ratio was 2.78:1. Pre-existing CVDs were present in 258 patients (50.5%). The most common cardiovascular manifestation was acute coronary syndrome (ACS), seen in 259 patients (50.7%). ST-segment elevation myocardial infarction (STEMI) was more common than non-ST-segment elevation ACS (NSTE-ACS). Possible myocarditis was seen in 52 patients (10.1%). Rhythm and conduction abnormalities were noted in 144 patients (28.2%), the most common being QT prolongation, seen in 51 patients (10%). In-hospital mortality occurred in 97 patients (18.9%). Age, serum Ferritin level, D-dimer, NT-pro-BNP and total leukocyte count were significantly higher among patients with in-hospital mortality compared to survival group. Blood lymphocyte count and Haemoglobin level were significantly lower in mortality-group, compared to survival-group. Incidence of pre-existing CVDs, cardiogenic shock, heart failure, atrial fibrillation (AF), and renal failure were significantly higher in mortality-group compared to survival-group.
The most common CVD in COVID-19 patients in our study was ACS. STEMI was more common than NSTE-ACS. Advanced age, elevated serum ferritin, D-dimer, NT pro-BNP, leucocytosis, lymphopenia, lower haemoglobin, pre-existing CVDs heart failure, cardiogenic shock, AF and renal failure were associated with increased mortality in these patients.
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