Phosphoproteome analysis of the crosstalk between sumoylation and phosphorylation in mouse spermatocytes.

Spermatogenesis is supported by various posttranslational modifications. There is growing evidence supporting a crosstalk between sumoylation and phosphorylation in different cell types. We have recently shown that inhibition of global sumoylation with a sumoylation inhibitor (Ginkgolic acid, GA) arrested purified mouse spermatocytes in vitro; the spermatocytes could not condense chromatin and disassemble the synaptonemal complex. Our data have also revealed that some kinases regulating the meiotic prophase (PLK1 and AURKB) were inhibited upon the inhibition of sumoylation. Nevertheless, specific phosphorylated targets affected by the inhibition of sumoylation have not been identified. To address this gap, in this study, we performed a comparative phospho-proteome analysis of the control spermatocytes and spermatocytes treated with the GA. Our analysis has narrowed down to several proteins implicated in the regulation of cell cycle and/or meiosis. Two of these targets, NPM1 and hnRNPH1, were studied further using western blotting in both cell lines and primary cells. Decrease in sumoylaion-dependend phosphorylation of NPM1 on Ser125 regulated by AURKB can be a contributing factor to the inability of spermatocytes to condense chromatin by the end of the prophase and should be studied further.

A Delphi consensus statement for the management of post-COVID interstitial lung disease.

INTRODUCTION: As millions of people worldwide recover from COVID-19, a substantial proportion continue to have persistent symptoms, pulmonary function abnormalities, and radiological findings suggestive of post-COVID interstitial lung disease (ILD). To date, there is limited scientific evidence on the management of post-COVID ILD, necessitating a consensus-based approach. AREAS COVERED: A panel of experts in pulmonology and thoracic radiology was constituted. Key questions regarding the management of post-COVID ILD were identified. A search was performed on PubMed and EMBASE and updated till 1 March 2022. The relevant literature regarding the epidemiology, pathophysiology, diagnosis and treatment of post-COVID ILD was summarized. Subsequently, suggestions regarding the management of these patients were framed, and a consensus was obtained using the Delphi approach. Those suggestions which were approved by over 80% of the panelists were accepted. The final document was approved by all panel members. EXPERT OPINION: Dedicated facilities should be established for the care of patients with post-COVID ILD. Symptom screening, pulmonary function testing, and thoracic imaging have a role in the diagnosis. The pharmacologic and non-pharmacologic options for the management of post-COVID ILD are discussed. Further research into the pathophysiology and management of post-COVID ILD will improve our understanding of this condition.

Clinical outcome of orbital apex syndrome in COVID associated mucormycosis patients in a tertiary care hospital.

AIM: To share clinical pattern of presentation, the modalities of surgical intervention and the one month post-surgical outcome of rhino-orbito-mucormycosis (ROCM) cases. METHODS: All COVID associated mucormycosis (CAM) patients underwent comprehensive multidisciplinary examination by ophthalmologist, otorhinolaryngologist and physician. Patients with clinical and radiological evidence of orbital apex involvement were included in the study. Appropriate medical and surgical intervention were done to each patient. Patients were followed up one-month post intervention. RESULTS: Out of 89 CAM patients, 31 (34.8%) had orbital apex syndrome. Sixty-six (74.2%) of such patients had pre-existing diabetes mellitus, 18 (58%) patients had prior documented use of steroid use, and 55 (61.8%) had no light perception (LP) presenting vision. Blepharoptosis, proptosis, complete ophthalmoplegia were common clinical findings. Seventeen (19.1%) of such patients had variable amount of cavernous sinus involvement. Endoscopic debridement of paranasal sinuses and orbit with or without eyelid sparing limited orbital exenteration was done in most cases, 34 (38.2%) patients could retain vision in the affected eye. CONCLUSION: Orbital apex involvement in CAM patients occur very fast. It not only leads to loss of vision but also sacrifice of the eyeball, orbital contents and eyelids. Early diagnosis and prompt intervention can preserve life, vision and spare mutilating surgeries.

Sumoylation and its regulation in testicular Sertoli cells.

The molecular regulation of Sertoli cells and their crosstalk with germ cells has not been fully characterized. SUMO proteins are essential for normal development and are expressed in mouse and human Sertoli cells; However, the cell-specific role of sumoylation in those cells has only started to be elucidated. In other cell types, including granulosa cells, sumoylation is regulated by a SUMO ligase KAP1/Trim28. Deletion of KAP1 in Sertoli cells causes testicular degeneration; However, the role of KAP1 in those cells has not been identified. Here we show that both mouse and human Sertoli undergo apoptosis upon inhibition of sumoylation with a chemical inhibitor or via a siRNA technology. We have additionally detected changes in the Sertoli cell proteome upon the inhibition of sumoylation, and our data suggest that among others, the expression of ER/stress-related proteins is highly affected by this inhibition. Sumoylation may also regulate the NOTCH signaling which is important for the maintenance of the developing germ cells. Furthermore, we show that a siRNA-down-regulation of KAP1 in a Sertoli-derived cell line causes an almost complete inactivation of sumoylation. In conclusion, sumoylation regulates important survival and signaling pathways in Sertoli cells, and KAP1 can be a major regulator of sumoylation in these cells.

COVID-19 associated rhino-orbital-cerebral mucormycosis: An observational study from Eastern India, with special emphasis on neurological spectrum.

AIMS: 1: Describe the epidemiology and determine risk factors for COVID-19 associated mucormycosis. 2: Elaborate the clinical spectrum of Rhino-Orbital-Cerebral Mucormycosis (ROCM), pattern of neuroaxis involvement and it's radiological correlates. METHODS: Observational study. Consecutive, confirmed cases of mucormycosis (N = 55) were included. A case of mucormycosis was defined as one who had clinical and radiological features consistent with mucormycosis along with demonstration of the fungus in tissue via KOH mount/culture/histopathological examination (HPE). Data pertaining to epidemiology, risk factors, clinico-radiological features were analysed using percentage of total cases. RESULTS: Middle aged, diabetic males with recent COVID-19 infection were most affected. New onset upper jaw toothache was a striking observation in several cases. Among neurological manifestations headache, proptosis, vision loss, extraocular movement restriction; cavernous sinus, meningeal and parenchymal involvement were common. Stroke in ROCM followed a definitive pattern with watershed infarction. CONCLUSIONS: New onset upper jaw toothache and loosening of teeth should prompt an immediate search for mucormycosis in backdrop of diabetic patients with recent COVID-19 disease, aiding earlier diagnosis and treatment initiation. Neuroaxis involvement was characterized by a multitude of features pertaining to involvement of optic nerve, extraocular muscles, meninges, brain parenchyma and internal carotid artery.

A rare case of isolated testicular tuberculosis and review of literature.

Testicular tuberculosis (TB) is a rare form of genitourinary TB. It is usually presented as painful or painless testicular swelling with or without scrotal ulceration or discharging sinus. Infertility may occur. Epididymal involvement is usually seen in testicular TB. In most cases, genital TB is associated with TB involvement of kidneys or lower urinary tract. Ultrasound (USG) and USG-guided fine needle aspiration cytology of testicular swelling confirm the diagnosis. Anti-TB chemotherapy is the mainstay of treatment to ensure the complete resolution of the lesion. However, in very few cases, orchidectomy is required for both diagnosis and treatment. Here, we report a very rare case of left sided isolated testicular TB in a 20-year-old male who was completely cured with 6 months regimen of anti-TB chemotherapy.

Staufen1 promotes HCV replication by inhibiting protein kinase R and transporting viral RNA to the site of translation and replication in the cells.

Persistent hepatitis C virus (HCV) infection leads to chronic hepatitis C (CHC), which often progresses to liver cirrhosis (LC) and hepatocellular carcinoma (HCC). The molecular mechanisms that establish CHC and cause its subsequent development into LC and HCC are poorly understood. We have identified a cytoplasmic double-stranded RNA binding protein, Stau1, which is crucial for HCV replication. In this study, Stau1 specifically interacted with the variable-stem-loop region in the 3' NTR and domain IIId of the HCV-IRES in the 5' NTR, and promoted HCV replication and translation. Stau1 coimmunoprecipitates HCV NS5B and a cell factor, protein kinase R (PKR), which is critical for interferon-induced cellular antiviral and antiproliferative responses. Like Stau1, PKR displayed binding specificity to domain IIId of HCV-IRES. Stau1 binds to PKR and strongly inhibits PKR-autophosphorylation. We demonstrated that the transport of HCV RNA on the polysomes is Stau1-dependent, being mainly localized in the monosome fractions when Stau1 is downregulated and exclusively localized in the polysomes when Stau1 is overexpressed. Our findings suggest that HCV may appropriate Stau1 to its advantage to prevent PKR-mediated inhibition of eIF2α, which is required for the synthesis of HCV proteins for translocation of viral RNA genome to the polysomes for efficient translation and replication.

Substrate Stiffness Regulates Proinflammatory Mediator Production through TLR4 Activity in Macrophages.

Clinical data show that disease adversely affects tissue elasticity or stiffness. While macrophage activity plays a critical role in driving disease pathology, there are limited data available on the effects of tissue stiffness on macrophage activity. In this study, the effects of substrate stiffness on inflammatory mediator production by macrophages were investigated. Bone marrow-derived macrophages were grown on polyacrylamide gels that mimicked the stiffness of a variety of soft biological tissues. Overall, macrophages grown on soft substrates produced less proinflammatory mediators than macrophages grown on stiff substrates when the endotoxin LPS was added to media. In addition, the pathways involved in stiffness-regulated proinflammation were investigated. The TLR4 signaling pathway was examined by evaluating TLR4, p-NF-κB p65, MyD88, and p-IκBα expression as well as p-NF-κB p65 translocation. Expression and translocation of the various signaling molecules were higher in macrophages grown on stiff substrates than on soft substrates. Furthermore, TLR4 knockout experiments showed that TLR4 activity enhanced proinflammation on stiff substrates. In conclusion, these results suggest that proinflammatory mediator production initiated by TLR4 is mechanically regulated in macrophages.

Unexplained dyspnea in a patient of chronic arsenicosis: A diagnostic challenge and learning curve for physicians.

Chronic arsenic exposure causes cutaneous effects like hyperkeratosis, peripheral vascular disease, hypertension, ischemic heart disease, non-cirrhotic portal hypertension, hepatomegaly, peripheral neuropathy, respiratory involvement, bad obstetrical outcome, hematological disturbances, and diabetes mellitus. Here we present a case of a 24-year-old lady, with chronic exposure to arsenic, presenting to us with progressive dyspnea. We found pulmonary arterial hypertension (PAH) as a cause of her dyspnea. PAH can occur in arsenicosis, secondary to arsenic-induced chronic obstructive pulmonary disease (COPD), lung fibrosis, and portal hypertension, which we excluded by appropriate investigations in our case. We also excluded a familial or heritable form of PAH. Thus, with the exclusion of all these secondary causes of PAH, as well as a hereditary cause, we came to a conclusion that this PAH might be due to chronic arsenic exposure. To the best of our knowledge, no case of PAH in chronic arsenicosis has been reported to date.

Role of DR-70 immunoassay in suspected malignant pleural effusion.

CONTEXT: A good proportion of patients with undiagnosed pleural effusion (PE) turn into malignancy over a period of time. Identification of positive biomarker may help in selecting the individuals who require close follow-up. AIMS: The aims of this study were to evaluate the role of DR-70 immunoassay in suspected malignant PE. SETTINGS AND DESIGN: We conducted a cross-sectional study among 89 patients of suspected malignant PE and 50 normal subjects (NS) were taken as control. MATERIALS AND METHODS: Patients with exudative PE; who had pleural fluid lymphocyte count greater than 50% and adenosine deaminase less than 30 U/L were taken as cases. We had selected NSs among relatives of patients having normal blood chemistry and radiological investigations. Sensitivity and specificity of the test to differentiate malignant and non-malignant PE and also to identify PE with underlying malignancy was analyzed. RESULTS: Mean value of DR-70 in NS was found to be 0.83 ± 0.273 mg/L without any significant difference between males (0.82 mg/L) and females (0.85 mg/L). Mean value of DR-70 in PE with underlying cancer was 5.03 ± 3.79 mg/L. Sensitivity (80%) and specificity (77.78%) of the test was maximum in PE with underlying cancer using cut-off value of 2 mg/L. Mean value DR-70 in malignant PE was 5.18 ± 3.75 mg/L and in non-malignant PE was 3.73 ± 3.74 mg/L without any statistically significant difference (P = 0.08). CONCLUSIONS: DR-70 assay has high sensitivity in detecting underlying lung cancer, but has no role in differentiating malignant PE from non-malignant PE.

Habitual physical activity score as a predictor of the 6-min walk test distance in healthy adults.

BACKGROUND: The 6-min walk test (6MWT) is a simple, inexpensive test of functional exercise capacity. The 6MWT distance (6MWD) in healthy adults varies geographically, emphasizing the need for population-specific reference equations. The purpose of the present study was to investigate the influences of the habitual physical activity (HPA) score and other anthropometric and demographic parameters on the variability of the 6MWD among healthy adults and to propose a reference equation. METHODS: This was a prospective, cross-sectional, observational study. The 6MWT was conducted in a 30-m hospital corridor on 201 healthy volunteers, 125 men and 76 women, aged 20-60 years. The HPA in the previous 6 months was assessed using Baecke's questionnaire. Univariate analysis followed by multiple regression analysis was performed to analyze the significance levels of different probable predictors. RESULTS: The 6MWD was significantly greater in more active than in less active subjects (663.8±55.4m vs. 599.9±67.8m, p<0.001). The regression analysis showed that the subject's age in years (p=0.017), gender (p=0.006), height in cm (p=0.004), weight in kg (p<0.001), total activity score (TS) (p<0.001), and absolute difference in heart rate before and after exercise (p<0.001) could explain 48.9% of the variability in the 6MWD in healthy adults. CONCLUSIONS: The HPA score is probably the most appropriate variable to include in the reference equation predicting the 6MWD in healthy adults from the Indian subcontinent.

A rare case of ethambutol induced pulmonary eosinophilia.

Antitubercular drug (ATD) induced eosinophilic lung disease is a rare phenomenon. It usually occurs due to isoniazid and para amino salicylic acid. A 34-year-male of sputum positive pulmonary tuberculosis, on antitubercular drugs (rifampicin, isoniazid, ethambutol, and pyrazinamide) for last 3 weeks, presented with generalized arthralgia and maculopapular rash for last 2 weeks and shortness of breath for last 1 week. Chest X-ray and High resolution computerized tomographic scan thorax showed bilateral peripheral airspace opacification. Bronchoalveolar lavage revealed 51% eosinophils of total cellularity (1200/cmm) confirming the diagnosis of pulmonary eosinophilia. ATD was stopped for 2 weeks and then reintroduced one by one. Patient again developed similar kind of symptoms with reintroduction of ethambutol. According to criteria for drug induced pulmonary eosinophilia, he was diagnosed as a case of ethambutol induced pulmonary eosinophilia.

The Incidence of Hyponatraemia and Its Effect on the ECOG Performance Status among Lung Cancer Patients.

CONTEXT: Hyponatraemia is one of the common electrolytic disorders which are associated with lung cancer. Hyponatraemia may influence the ECOG performance status at presentation. Also, to the best of our knowledge, we found only limited Indian studies where the ECOG score was correlated with the serum sodium status in lung cancer patients on presentation. AIM: To assess the incidence of hyponatraemia among the patients of carcinoma of the lung before putting them into the specific treatment category for cancer and to check the effects on their ECOG performance status. SETTINGS AND DESIGN: A cross-sectional, observational study was conducted on 116 consecutive patients of lung cancer during the period from November 2011 to October 2012. MATERIAL AND METHODS: The patients with a histologically proven diagnosis of lung cancer were grouped initially according to their ECOG performance statuses. The serum sodium value of each patient was measured and the hyponatraemic patients were given treatment according to the protocol. The correlation of the ECOG performance status with the serum sodium of the lung cancer patients was measured. To check for any laboratory error in serum sodium, we selected (n = 58) age, sex and socioeconomic matched control patients. RESULTS: At presentation 44.8% of the lung cancer patients showed hyponatraemia [52/116]. The ECOG score was significantly poor in the advanced clinical stages (ECOG ≤2 Vs ECOG ≥ 3 in NSCLC cases, χ(2) =11.25, P=.0008). The ECOG performance status score at admission showed a negative correlation with the serum sodium status which was measured on admission among all the patients (Pearson correlation coefficient = - 0.186). The clinical stage of the lung cancer also showed a positive correlation with the ECOG score at admission in our study (Pearson correlation coefficient = 0.295). CONCLUSION: Hyponatraemia is not an uncommon condition and it should be suspected and screened in each patient, as it may influence the ECOG performance status score, which serves as an important factor in the prognosis of lung cancer.

Dyspnea with anemia turned out to be a case of hereditary hemorrhagic telangiectasia.

Hereditary hemorrhagic telangiectasia (HHT) is a rare autosomal dominant inherited disorder of the vascular system. It can be asymptomatic but when symptomatic most common presentation being epistaxis. It can involve any organs of the body like lungs, skin, liver brain, GI mucosa etc. We are reporting a case of HHT presented to us with dyspnea and severe anemia. He had arteriovenous malformations of different visceral organs and telangiectasia of skin along with presence of similar history in first-degree relatives.

Squamous cell lung cancer producing bilateral air bronchograms on CT scan thorax.

A young lady presented with cough for three months and dyspnoea for one month along with clinical signs of bilateral consolidation of lung. On CT scan thorax there were bilateral air bronchograms with alveolar filling opacities predominantly in lower lobes. After bronchoscopy when there was presence of suspicious malignant cells on bronchoalveolar lavage fluid, transbronchial lung biopsy diagnosed the case as squamous cell lung cancer. Radiological presentation of squamous cell lung cancer as air bronchogram is not a very common phenomenon and can present a diagnostic challenge to the clinician.

Squamous cell carcinoma lung: Presented with bilateral lower limb deep venous thrombosis with gangrene formation.

Bilateral venous thrombosis due to underlying malignancy is a rare entity. It is worthy to search for malignancy in patients of bilateral venous gangrene. Our patient presented with severe bilateral leg pain as a result of venous gangrene. There was associated left sided massive pleural effusion with scalp nodule. Fine needle aspiration cytology of scalp nodule revealed metastatic squamous cell carcinoma and fiber optic bronchoscopy guided biopsy from growth at left upper lobe bronchus confirmed the case as squamous cell carcinoma lung. It was rare for squamous cell carcinoma lung to present as bilateral venous gangrene with anticardiolipin antibody negative.

Roles of dopamine 2 receptor isoforms and g proteins in ethanol regulated prolactin synthesis and lactotropic cell proliferation.

Alcohol consumption has been shown to increase prolactin (PRL) production and cell proliferation of pituitary lactotropes. It also causes a reduction in the lactotrope's response to dopaminergic agents and a differential expression of dopamine 2 receptor short (D2S) and long (D2L) isoforms in the pituitary. However, the role of each of these D2 receptor isoforms and its coupled G protein in mediation of ethanol actions on lactotropes is not known. We have addressed this issue by comparing ethanol effects on the level of PRL production gene transcription rate cellular protein, G proteins and cell proliferation in enriched lactotropes and lactotrope-derived PR1 cells containing various D2 receptor isoforms. Additionally, we determined the effects of G protein blockade on ethanol-induced PRL production and cell proliferation in these cells. We show here that the D2 receptor, primarily the D2S isoform, is critically involved in the regulation of ethanol actions on PRL production and cell proliferation in lactotropes. We also present data to elucidate that the presence of the pertussis toxin (PTX)-sensitive D2S receptor is critical to mediate the ethanol stimulatory action on Gs and the ethanol's inhibitory action on Gi3 protein in lactotropes. Additionally, we provide evidence for the existence of an inhibitory action of Gi3 on Gs that is under the control of the D2S receptor and is inhibited by ethanol. These results suggest that ethanol via the inhibitory action on D2S receptor activity suppresses Gi3 repression of Gs expression resulting in stimulation of PRL synthesis and cell proliferation in lactotropes.

Opiate antagonist prevents µ- and δ-opiate receptor dimerization to facilitate ability of agonist to control ethanol-altered natural killer cell functions and mammary tumor growth.

In the natural killer (NK) cells, δ-opiate receptor (DOR) and µ-opioid receptor (MOR) interact in a feedback manner to regulate cytolytic function with an unknown mechanism. Using RNK16 cells, a rat NK cell line, we show that MOR and DOR monomer and dimer proteins existed in these cells and that chronic treatment with a receptor antagonist reduced protein levels of the targeted receptor but increased levels of opposing receptor monomer and homodimer. The opposing receptor-enhancing effects of MOR and DOR antagonists were abolished following receptor gene knockdown by siRNA. Ethanol treatment increased MOR and DOR heterodimers while it decreased the cellular levels of MOR and DOR monomers and homodimers. The opioid receptor homodimerization was associated with an increased receptor binding, and heterodimerization was associated with a decreased receptor binding and the production of cytotoxic factors. Similarly, in vivo, opioid receptor dimerization, ligand binding of receptors, and cell function in immune cells were promoted by chronic treatment with an opiate antagonist but suppressed by chronic ethanol feeding. Additionally, a combined treatment of an MOR antagonist and a DOR agonist was able to reverse the immune suppressive effect of ethanol and reduce the growth and progression of mammary tumors in rats. These data identify a role of receptor dimerization in the mechanism of DOR and MOR feedback interaction in NK cells, and they further elucidate the potential for the use of a combined opioid antagonist and agonist therapy for the treatment of immune incompetence and cancer and alcohol-related diseases.