The Role of Aging and Senescence in Immune Checkpoint Inhibitor Response and Toxicity.

Cellular senescence accumulates with age and has been shown to impact numerous physiological and pathological processes, including immune function. The role of cellular senescence in cancer is multifaceted, but the impact on immune checkpoint inhibitor response and toxicity has not been fully evaluated. In this review, we evaluate the impact of cellular senescence in various biological compartments, including the tumor, the tumor microenvironment, and the immune system, on immune checkpoint inhibitor efficacy and toxicity. We provide an overview of the impact of cellular senescence in normal and pathological contexts and examine recent studies that have connected aging and cellular senescence to immune checkpoint inhibitor treatment in both the pre-clinical and clinical contexts. Overall, senescence plays a multi-faceted, context-specific role and has been shown to modulate immune-related adverse event incidence as well as immune checkpoint inhibitor response.

Circulating, cell-free methylated DNA indicates cellular sources of allograft injury after liver transplant.

Post-transplant complications reduce allograft and recipient survival. Current approaches for detecting allograft injury non-invasively are limited and do not differentiate between cellular mechanisms. Here, we monitor cellular damages after liver transplants from cell-free DNA (cfDNA) fragments released from dying cells into the circulation. We analyzed 130 blood samples collected from 44 patients at different time points after transplant. Sequence-based methylation of cfDNA fragments were mapped to patterns established to identify cell types in different organs. For liver cell types DNA methylation patterns and multi-omic data integration show distinct enrichment in open chromatin and regulatory regions functionally important for the respective cell types. We find that multi-tissue cellular damages post-transplant recover in patients without allograft injury during the first post-operative week. However, sustained elevation of hepatocyte and biliary epithelial cfDNA beyond the first week indicates early-onset allograft injury. Further, cfDNA composition differentiates amongst causes of allograft injury indicating the potential for non-invasive monitoring and timely intervention.

Snake Bite-Induced Leukoencephalopathy: A Rare Case.

Across the globe, snake envenomation causes significant morbidity and mortality. Although many clinical presentations and complications are observed in different types of snake bites, the incidence of leukoencephalopathy is rare. Although most cases of leukoencephalopathy are seen in viper bites, they are rarely seen in neurotoxic snake bites. In this report, we present a unique case of snake bite-induced leukoencephalopathy following a neurotoxic snake bite. The case highlights the importance of considering this rare complication in cases of snake bites presenting with neurological symptoms, particularly in those affecting higher mental functions.

A ketogenic diet reduces age-induced chronic neuroinflammation in mice Running title: ketogenic diet and brain inflammaging.

Beta-hydroxybutyrate (BHB) is a ketone body synthesized during fasting or strenuous exercise. Our previous study demonstrated that a cyclic ketogenic diet (KD), which induces BHB levels similar to fasting every other week, reduces midlife mortality and improves memory in aging mice. BHB actively regulates gene expression and inflammatory activation through non-energetic signaling pathways. Neither of these activities has been well-characterized in the brain and they may represent mechanisms by which BHB affects brain function during aging. First, we analyzed hepatic gene expression in an aging KD-treated mouse cohort using bulk RNA-seq. In addition to the downregulation of TOR pathway activity, cyclic KD reduces inflammatory gene expression in the liver. We observed via flow cytometry that KD also modulates age-related systemic T cell functions. Next, we investigated whether BHB affects brain cells transcriptionally in vitro. Gene expression analysis in primary human brain cells (microglia, astrocytes, neurons) using RNA-seq shows that BHB causes a mild level of inflammation in all three cell types. However, BHB inhibits the more pronounced LPS-induced inflammatory gene activation in microglia. Furthermore, we confirmed that BHB similarly reduces LPS-induced inflammation in primary mouse microglia and bone marrow-derived macrophages (BMDMs). BHB is recognized as an inhibitor of histone deacetylase (HDAC), an inhibitor of NLRP3 inflammasome, and an agonist of the GPCR Hcar2. Nevertheless, in microglia, BHB's anti-inflammatory effects are independent of these known mechanisms. Finally, we examined the brain gene expression of 12-month-old male mice fed with one-week and one-year cyclic KD. While a one-week KD increases inflammatory signaling, a one-year cyclic KD reduces neuroinflammation induced by aging. In summary, our findings demonstrate that BHB mitigates the microglial response to inflammatory stimuli, like LPS, possibly leading to decreased chronic inflammation in the brain after long-term KD treatment in aging mice.

A visible light-driven direct synthesis of industrially relevant glutaric acid diesters from aldehydes.

A straightforward and practical method has been developed to access α-substituted glutaric diesters from acrylates and aldehydes using visible light, with Eosin Y facilitating hydrogen atom transfer (HAT) and subsequent Giese-type addition. Also, sunlight has been successfully used as an alternative sustainable light source. The method has also been explored to access substituted 4,5-dihydro-2H-pyridazinones, which have potential biological and industrial applications. Comprehensive mechanistic investigations have been carried out.

Deconvolution of immune cell composition and biological age of hepatocellular carcinoma using DNA methylation.

Hepatocellular carcinoma (HCC) has been an approved indication for the administration of immunotherapy since 2017, but biomarkers that predict therapeutic response have remained limited. Understanding and characterizing the tumor immune microenvironment enables better classification of these tumors and may reveal biomarkers that predict immunotherapeutic efficacy. In this paper, we applied a cell-type deconvolution algorithm using DNA methylation array data to investigate the composition of the tumor microenvironment in HCC. Using publicly available and in-house datasets with a total cohort size of 57 patients, each with tumor and matched normal tissue samples, we identified key differences in immune cell composition. We found that NK cell abundance was significantly decreased in HCC tumors compared to adjacent normal tissue. We also applied DNA methylation "clocks" which estimate phenotypic aging and compared these findings to expression-based determinations of cellular senescence. Senescence and epigenetic aging were significantly increased in HCC tumors, and the degree of age acceleration and senescence was strongly associated with decreased NK cell abundance. In summary, we found that NK cell infiltration in the tumor microenvironment is significantly diminished, and that this loss of NK abundance is strongly associated with increased senescence and age-related phenotype. These findings point to key interactions between NK cells and the senescent tumor microenvironment and offer insights into the pathogenesis of HCC as well as potential biomarkers of therapeutic efficacy.

Circulating cell-free methylated DNA reveals tissue-specific, cellular damage from radiation treatment.

Radiation therapy is an effective cancer treatment, although damage to healthy tissues is common. Here we analyzed cell-free, methylated DNA released from dying cells into the circulation to evaluate radiation-induced cellular damage in different tissues. To map the circulating DNA fragments to human and mouse tissues, we established sequencing-based, cell-type-specific reference DNA methylation atlases. We found that cell-type-specific DNA blocks were mostly hypomethylated and located within signature genes of cellular identity. Cell-free DNA fragments were captured from serum samples by hybridization to CpG-rich DNA panels and mapped to the DNA methylation atlases. In a mouse model, thoracic radiation-induced tissue damage was reflected by dose-dependent increases in lung endothelial and cardiomyocyte methylated DNA in serum. The analysis of serum samples from patients with breast cancer undergoing radiation treatment revealed distinct dose-dependent and tissue-specific epithelial and endothelial responses to radiation across multiple organs. Strikingly, patients treated for right-sided breast cancers also showed increased hepatocyte and liver endothelial DNA in the circulation, indicating the impact on liver tissues. Thus, changes in cell-free methylated DNA can uncover cell-type-specific effects of radiation and provide a readout of the biologically effective radiation dose received by healthy tissues.

Light-cured calcium hydroxide cements release of calcium ions using argon based induction coupled mass spectroscopy - an in vitro study.

Calcium ion-releasing ability of different calcium hydroxide-based pulp capping materials was comparatively evaluated in this study. Different brands of cements were taken from different manufacturers and categorized into three groups. Three different brands of Ca(OH)2 cements (Dycal, TheraCal, and Cal LC) were taken prepared by mixing and curing the cements as per the manufacturer's instructions. Consequently, ion release was measured after 7, 14, and 21 days by argon-based induction coupled plasma mass spectroscopy test. Within the limitations of this study, light-cured Ca(OH)2 cements released a higher amount of calcium ions compared with self-cured Ca(OH)2 cements. Theracal was found to be the highest light-cured calcium ion releasing materials throughout the period of 21 days. In conclusion, further clinical studies are warranted to substantiate the findings of this study.

Hexameric Aggregation Nucleation Core Sequences and Diversity of Pathogenic Tau Strains.

Tau aggregation associates with multiple neurodegenerative diseases including Alzheimer's disease and rare tauopathies such as Pick's disease, progressive supranuclear palsy, and corticobasal degeneration. The molecular and structural basis of tau aggregation and related diverse misfolded tau strains are not fully understood. To further understand tau-protein aggregation mechanisms, we performed systematic truncation mutagenesis and mapped key segments of tau proteins that contribute to tau aggregation, where it was determined that microtubule binding domains R2 and R3 play critical roles. We validated that R2- or R3-related hexameric PHF6 and PHF6* peptide sequences are necessary sequences that render tau amyloidogenicity. We also determined that the consensus VQI peptide sequence is not sufficient for amyloidogenicity. We further proposed single- and dual-nucleation core-based strain classifications based on recent cryo-EM structures. We analyzed the structural environment of the hexameric peptide sequences in diverse tau strains in tauopathies that, in part, explains why the VQI consensus core sequence is not sufficient to induce tau aggregation. Our experimental work and complementary structural analysis highlighted the indispensible roles of the hexameric core sequences, and shed light on how the interaction environment of these core sequences contributes to diverse pathogenic tau-strains formation in various tauopathy brains.

Role of Ambulatory Glucose Profile in Precision Medicine in Type 2 Diabetes Mellitus.

Assessment of diabetes with daily blood glucose fluctuations including peaks and nadirs forms the crux of the modern management. Use of glycemic variability (GV) as a parameter to assess these fluctuations is emerging. It is important to determine the hyperglycemic and hypoglycemic episodes which are the culprits for increasing glycemic variation. Diabetes mellitus patients follow different clinical trajectories which can be traced by the ambulatory glucose profile (AGP) obtained from flash glucose monitoring system (FGMS). MATERIAL: This comparative observational study enrolled 106 adult (>18 years) type 2 diabetes patients with HbA1c<8%. Patients were divided into two groups (group A & group B) with 53 patients each. Group A included patients on OAD's (oral antidiabetic drug) with insulin and Group B included patients on OAD's without insulin. The patients were put on FGMS for 14 days and their AGP was analysed. Hyperglycemic episodes (level 1- >180 mg/dl, level 2- >250 mg/dl) and hypoglycemic episodes (level 1- 54-70 mg/dl, level 2- <54 mg/dl) were determined between the groups. OBSERVATION: Group A patients had significantly higher (29.99%) total number of hyperglycemic episodes (Level 1+ Level 2) as compared with group B (9.08%) (p <0.0001). Amongst group A, proportion of patients with total number of hyperglycemic episodes was significantly higher in insulin only subgroup (58.11%) followed by insulin +metformin+ 1 OAD (29.14%) & insulin+ metformin (26.82%) (p <0.0001). Amongst group B, total number of hyperglycemic episodes were found to be significantly higher with metformin only subgroup (10.19%) followed by metformin + 1 OAD (9.72%) & metformin + >1 OAD (8.1%) (p<0.0001). Amongst the add on OAD's, sulfonylurea contributed to 61.07% hyperglycemic episodes in group A & 11.63% in group B which was statistically more than DPP-4 inhibitors with 14.91% & 2.84% respectively (p <0.0001). Total number of hypoglycemic episodes seen in group A patients (8.66%) were significantly less as compared with group B (13.27%) (p<0.0001). Sulfonylurea contributed to 7.5% hypoglycemic episodes in group A & 13.2% in group B which was statistically more than DPP-4 inhibitors with 6.49% & 12.35% respectively when added to metformin (p<0.0001). CONCLUSION: Amongst the OAD's used in type 2 diabetes mellitus patients in this study, total number of hyperglycemic and hypoglycemic episodes were found to be more in patients taking sulfonylurea as compared with DPP4 inhibitors when used in combination with metformin with or without insulin.

A case of Weil's disease presenting as Stevens-Johnson syndrome.

Leptospirosis presents in a biphasic manner: an early leptospiraemic phase and a late immune phase. In its severe form, it presents with multi-organ failure, also known as Weil's disease. Stevens-Johnson syndrome (SJS) is an autoimmune hypersensitive reaction leading to diffuse fluid filled vesicle formation with detachment of skin and mucous membrane. Though SJS is triggered by different infections and drugs, its association with leptospirosis is not frequently reported. Here we present such a case.

Cell-type Deconvolution and Age Estimation Using DNA Methylation Reveals NK Cell Deficiency in the Hepatocellular Carcinoma Microenvironment.

Hepatocellular carcinoma (HCC) has been an approved indication for the administration of immunotherapy since 2017, but biomarkers that predict therapeutic response have remained limited. Understanding and characterizing the tumor immune microenvironment enables better classification of these tumors and may reveal biomarkers that predict immunotherapeutic efficacy. In this paper, we applied a cell-type deconvolution algorithm using DNA methylation array data to investigate the composition of the tumor microenvironment in HCC. Using two publicly available datasets with a total cohort size of 57 patients, each with tumor and matched normal tissue samples, we identified key differences in immune cell composition. We found that NK cell abundance was significantly decreased in HCC tumors compared to adjacent normal tissue. We also applied DNA methylation "clocks" which estimate phenotypic aging and compared these findings to expression-based determinations of cellular senescence. Senescence and epigenetic aging was significantly increased in HCC tumors, and the degree of age acceleration and senescence was strongly associated with decreased NK cell abundance. In summary, we found that NK cell infiltration in the tumor microenvironment is significantly diminished, and that this loss of NK abundance is strongly associated with increased senescence and age-related phenotype. These findings point to key interactions between NK cells and the senescent tumor microenvironment and offer insights into the pathogenesis of HCC as well as potential biomarkers of therapeutic efficacy.

Solid Organ Abscess by Melioidosis: An Emerging Infection Among Diabetics in Odisha, India.

Melioidosis is an infection in tropical regions, caused by Gram-negative bacillus Burkholderia pseudomallei and carries very high case fatality rates. Diabetes mellitus is a major risk factor for melioidosis, and Odisha with its current trends in urban prevalence of diabetes and a tropical coastal climate seems to be emerging as a hub of melioidosis. We herein describe two cases of melioidosis encountered within a month. Case 1 is a 35-year-old male recently diagnosed with diabetes, presenting with fever and altered sensorium for 5 days. Case 2 is a 40-year-old female who complained of fever, cough, and generalised weakness for 2 months, and was subsequently found to be diabetic on routine investigations. Both the patients were subjected to imaging modalities which showed solid organ (liver and spleen) abscesses. On further investigations, Burkholderia pseudomallei was isolated from their blood cultures. Diagnosis of such cases within a short period of time points to the fact that Odisha indeed serves as a hidden hotspot of melioidosis. The case report throws light on the importance of early diagnosis and treatment of melioidosis, which is under-reported in the state, and it also aims to impress upon physicians to have a strong clinical suspicion so as to decrease the mortality associated this neglected tropical disease.

Molecular basis for substrate recruitment to the PRMT5 methylosome.

PRMT5 is an essential arginine methyltransferase and a therapeutic target in MTAP-null cancers. PRMT5 uses adaptor proteins for substrate recruitment through a previously undefined mechanism. Here, we identify an evolutionarily conserved peptide sequence shared among the three known substrate adaptors (CLNS1A, RIOK1, and COPR5) and show that it is necessary and sufficient for interaction with PRMT5. We demonstrate that PRMT5 uses modular adaptor proteins containing a common binding motif for substrate recruitment, comparable with other enzyme classes such as kinases and E3 ligases. We structurally resolve the interface with PRMT5 and show via genetic perturbation that it is required for methylation of adaptor-recruited substrates including the spliceosome, histones, and ribosomal complexes. Furthermore, disruption of this site affects Sm spliceosome activity, leading to intron retention. Genetic disruption of the PRMT5-substrate adaptor interface impairs growth of MTAP-null tumor cells and is thus a site for development of therapeutic inhibitors of PRMT5.

When are we Going to Settle the Diagnostic Criteria of Gestational Diabetes Mellitus?

Guidelines to diagnose Gestational Diabetes Mellitus (GDM) have changed a number of times from O'Sullivan and Mahan, Carpenter and Coustan, World Health Organization, American Diabetes Association to that of International Association of Diabetes in Pregnancy Study Group (IADPSG). The IADPSG guideline was based on Hyperglycaemia and Adverse Pregnancy Outcome (HAPO) study which was performed in caucasian population only and thus literally cannot be considered as international. Recently a study commented that this guideline needs revision for standardization of this strategy for diagnosing GDM. Based on a prospective study, Diabetes in Pregnancy Study Group India (DIPSI) recommended A single step procedure of diagnosing GDM with 2hr PG > 140 mg/dl after 75g of oral glucose administered irrespective of the last meal timing. This guideline has been approved by the Ministry of Health Government of India, WHO, IDF and Federation of Gynaecologists and Obstetricians Society (FIGO). National Institute of Clinical Excellence (NICE) also recognises cut off value, 2hr PG > 140 mg/dl based on a study in multi ethnic population of UK. Hence, we can safely conclude, A Single Step procedure has settled the criteria for diagnosing GDM.

GEMINI: a variational Bayesian approach to identify genetic interactions from combinatorial CRISPR screens.

Systems for CRISPR-based combinatorial perturbation of two or more genes are emerging as powerful tools for uncovering genetic interactions. However, systematic identification of these relationships is complicated by sample, reagent, and biological variability. We develop a variational Bayes approach (GEMINI) that jointly analyzes all samples and reagents to identify genetic interactions in pairwise knockout screens. The improved accuracy and scalability of GEMINI enables the systematic analysis of combinatorial CRISPR knockout screens, regardless of design and dimension. GEMINI is available as an open source R package on GitHub at https://github.com/sellerslab/gemini .

Post-operative analgesia for shoulder arthroscopic surgeries: A comparison between inter-scalene block and shoulder block.

BACKGROUND AND AIMS: Shoulder arthroscopic surgeries can produce intense post-operative pain. Inter-scalene block (ISB) provides good analgesia after shoulder surgery, but concerns over its associated risks have prompted the search for alternatives. Shoulder block (SHB), which includes suprascapular block along with axillary nerve (AN) block, was recently proposed as an alternative to ISB, but evidence of its efficacy is conflicting. The aim of our study was to compare SHB with ISB in shoulder surgery for post-operative analgesia. METHODS: A total of 76 patients scheduled for shoulder arthroscopic surgery were equally divided into 2 groups of 38 patients each: ISB group and SHB group. Both the nerve blocks were achieved by using ultrasound and a nerve stimulator. Visual analogue scale (VAS) scores were evaluated at 1, 4, 6, 12 and 24 h post-operatively. The time to first analgesia request, total analgesic requirement for 24 h post-operatively, patient satisfaction and any complications were recorded. RESULTS: SHB provided equivalent analgesia to ISB in terms of post-operative VAS scores. Time to first analgesic request was 6.2 ± 1.3 h in ISB group and 5.9 ± 1.2 h in SHB group, which was not statistically significant. Complications like subjective dyspnoea and weakness of arm were significantly higher in ISB group compared to SHB group. Patient satisfaction scores were also significantly higher in SHB group compared to ISB group. CONCLUSION: SHB is as effective as ISB for post-operative pain relief and with fewer complications due to selective blockade of suprascapular and axillary nerves.

Clinical Characterization of Genetically Diagnosed Cases of Spinocerebellar Ataxia Type 12 from India.

https://onlinelibrary.wiley.com/page/journal/23301619/homepage/mdc312551-sup-v001_1.htm. BACKGROUND: Spinocerebellar ataxia type 12 (SCA12) is a rare form of an autosomal-dominant ataxic disorder associated with an expansion of CAG repeat length. Here, we present a large case series of patients with SCA12 and describe a wide range of typical and rare symptoms. METHODS: Twenty-one consecutive patients with genetically proven SCA12 underwent detailed neurological examination. We assessed clinical characteristics using validated rating scales for evaluating motor features in SCA. Nonmotor symptoms and quality of life were assessed using appropriate, validated scales. Correlations of CAG repeat length with both severity score and age of onset were explored. RESULTS: The mean age of onset was 51 years, and most patients were descendants of a single, endogamous Indian community (Agarwal). Tremor was the most common initial presenting symptom (90%). Hand dystonia was present in 14 of 21 patients, and most patients in the cohort presented with gait disturbance. Neuropsychiatric manifestations were common coexisting features. The CAG repeat length was significantly correlated (r = -0.760; P = 0.0001) with early age of onset, but not with disease severity. Tremor affected the quality of life in 18 of 21 patients, because they had difficulty in handling liquids. CONCLUSIONS: Tremor was the most common, nonataxic symptom at initial presentation in patients with SCA12. Proximal upper limb tremor, typically with high amplitude and low frequency, can raise a strong diagnostic suspicion. Associated hand dystonia was a common coexisting motor feature. Various nonmotor features were also observed in several cases which require therapeutic attention.

Comparison of pregabalin with gabapentin as preemptive analgesic in lumbar spine surgery.

BACKGROUND AND AIMS: Analgesic effect of gabapentin and pregabalin is well-defined in the treatment of neuropathic pain. Postoperative pain after lumbar spine surgery limits the function of patients in the postoperative period, for which the search for ideal analgesic goes on. The aim of the present study was to compare pregabalin and gabapentin as a pre-emptive analgesic in elective lumbar spine surgeries. MATERIAL AND METHODS: In this randomized prospective study, 75 patients were allocated into three groups of 25 each. Group G, group PG, and group P received two capsules of gabapentin 300 mg each, two capsules of pregabalin 150 mg each, and two multivitamin capsules, respectively, with sip of water 1 hour before the expected time of induction of anesthesia. Time for requirement of first dose of rescue analgesia, reduction in postoperative pain score and total dose of rescue analgesic used in first 24 hours postoperatively, and side effects were compared. RESULT: Time for requirement of first dose of rescue analgesic in PG group was 180.12 min and in G group was 104.16 min, which was statistically significant. Both G and PG group had lower visual analogue scale (VAS) score in comparison to P group, which was statistically significant. Consumption of rescue analgesic was less in G and PG group in comparison to P group. Amount of rescue analgesic requirement were low in PG group in comparison to G group (P < 0.001). CONCLUSION: Though both study drugs had produced prolonged postoperative analgesia compared to placebo, pregabalin had better analgesic profile in postoperative period than gabapentin.