Distinct metabolic responses to heme in inflammatory human and mouse macrophages - Role of nitric oxide.

Activation of inflammation is tightly associated with metabolic reprogramming in macrophages. The iron-containing tetrapyrrole heme can induce pro-oxidant and pro-inflammatory effects in murine macrophages, but has been associated with polarization towards an anti-inflammatory phenotype in human macrophages. In the current study, we compared the regulatory responses to heme and the prototypical Toll-like receptor (TLR)4 ligand lipopolysaccharide (LPS) in human and mouse macrophages with a particular focus on alterations of cellular bioenergetics. In human macrophages, bulk RNA-sequencing analysis indicated that heme led to an anti-inflammatory transcriptional profile, whereas LPS induced a classical pro-inflammatory gene response. Co-stimulation of heme with LPS caused opposing regulatory patterns of inflammatory activation and cellular bioenergetics in human and mouse macrophages. Specifically, in LPS-stimulated murine, but not human macrophages, heme led to a marked suppression of oxidative phosphorylation and an up-regulation of glycolysis. The species-specific alterations in cellular bioenergetics and inflammatory responses to heme were critically dependent on the availability of nitric oxide (NO) that is generated in inflammatory mouse, but not human macrophages. Accordingly, studies with an inducible nitric oxide synthase (iNOS) inhibitor in mouse, and a pharmacological NO donor in human macrophages, reveal that NO is responsible for the opposing effects of heme in these cells. Taken together, the current findings indicate that NO is critical for the immunomodulatory role of heme in macrophages.

Vaccine Breakthrough Infections Among Healthcare Workers in a COVID-19-Designated Tertiary Care Government Hospital in Sikkim.

Introduction Since the emergence of the coronavirus disease 2019 (COVID-19) virus at the beginning of 2020, the world has gone through various waves of pandemics. The health care workers (HCWs) or the COVID warriors as they were termed were the first line of defense against the virus. They were armed with personal protective equipment and prophylactic doses of the COVID-19 vaccine. Despite these precautions, some of the HCWs still contracted the disease and a few others succumbed to it. The objective of this study was to estimate the prevalence of COVID-19 infections and vaccine breakthrough infections (BTIs) in HCWs after receiving the COVID-19 vaccine during the second wave of the pandemic. Methods This was a cross-sectional, hospital-based study conducted over a period of four months from September 2021 to December 2021 on HCWs aged 18 years and above working at the COVID-19-designated tertiary care government hospital in Sikkim. A structured coded questionnaire with no patient identifiers was used to gather details on demographics, vaccination history, breakthrough infection, and other social details. HCWs who had received at least one dose of the COVID-19 vaccine at the time of initiation of the study and were >18 years of age were included in this study. Results A total of 678 HCWs were screened, out of which 229 (33%) participants tested positive for COVID-19 and the rest of the participants (455; 67%) tested negative. COVID-19 infections and vaccine BTIs (COVID-19 infection >14 days after the second vaccination) were recorded and 137 (20%) respondents had a post-vaccination COVID-19 infection out of which 115 (18.5%) were BTI. The majority of the participants were females and of the age group of 26-35 years. The correlation of COVID-19 infections with the dose gap between vaccination, gender, age, profession, department, area posted during COVID duty, cycles of duty performed, hospitalization due to infection, influenza vaccination, and comorbidity was analyzed. Conclusion COVID-19 vaccines are disease-modifying and they decrease the severity of BTIs in HCWs. Pandemics and outbreaks cannot be predicted; therefore, it becomes very important to have healthy frontline workers who are constantly exposed to infectious agents. Monitoring of health and surveillance of infectious diseases among the HCWs should be encouraged.

Using machine learning to study the effect of medication adherence in Opioid Use Disorder.

BACKGROUND: Opioid Use Disorder (OUD) and opioid overdose (OD) impose huge social and economic burdens on society and health care systems. Research suggests that Medication for Opioid Use Disorder (MOUD) is effective in the treatment of OUD. We use machine learning to investigate the association between patient's adherence to prescribed MOUD along with other risk factors in patients diagnosed with OUD and potential OD following the treatment. METHODS: We used longitudinal Medicaid claims for two selected US states to subset a total of 26,685 patients with OUD diagnosis and appropriate Medicaid coverage between 2015 and 2018. We considered patient age, sex, region level socio-economic data, past comorbidities, MOUD prescription type and other selected prescribed medications along with the Proportion of Days Covered (PDC) as a proxy for adherence to MOUD as predictive variables for our model, and overdose events as the dependent variable. We applied four different machine learning classifiers and compared their performance, focusing on the importance and effect of PDC as a variable. We also calculated results based on risk stratification, where our models separate high risk individuals from low risk, to assess usefulness in clinical decision-making. RESULTS: Among the selected classifiers, the XGBoost classifier has the highest AUC (0.77) closely followed by the Logistic Regression (LR). The LR has the best stratification result: patients in the top 10% of risk scores account for 35.37% of overdose events over the next 12 month observation period. PDC score calculated over the treatment window is one of the most important features, with better PDC lowering risk of OD, as expected. In terms of risk stratification results, of the 35.37% of overdose events that the predictive model could detect within the top 10% of risk scores, 72.3% of these cases were non-adherent in terms of their medication (PDC <0.8). Targeting the top 10% outcome of the predictive model could decrease the total number of OD events by 10.4%. CONCLUSIONS: The best performing models allow identification of, and focus on, those at high risk of opioid overdose. With MOUD being included for the first time as a factor of interest, and being identified as a significant factor, outreach activities related to MOUD can be targeted at those at highest risk.

Determination of free heme in stored red blood cells with an apo-horseradish peroxidase-based assay.

Transfusion effectiveness of red blood cells (RBCs) has been associated with duration of the storage period. Storage-dependent RBC alterations lead to hemolysis and release of toxic free heme, but the increase of free heme levels over time is largely unknown. In the current study, an apo-horseradish peroxidase (apoHRP)-based assay was applied to measure levels of free heme at regular intervals or periodically in supernatants of RBCs until a maximum storage period of 42 days. Free heme levels increased with linear time-dependent kinetics up to day 21 and accelerated disproportionally after day 28 until day 42, as determined with the apoHRP assay. Individual time courses of free heme in different RBC units exhibited high variability. Notably, levels of free hemoglobin, an established indicator of RBC damage, and those of total heme increased with continuous time-dependent linear kinetics over the entire 42 day storage period, respectively. Supernatants from RBC units with high levels of free heme led to inflammatory activation of human neutrophils. In conclusion, determining free heme in stored RBCs with the applied apoHRP assay may become feasible for testing of RBC storage quality in clinical transfusion medicine.

Improving access to quality family planning services in Nepal and Sri Lanka: insights from a South-South learning exchange.

Nepal and Sri Lanka ministries of health shared best practices and learnings, in a South-South learning exchange (SSLE) to improve access to quality and rights-based family planning services. The SSLE between the two countries followed a five-step methodology designed by the WHO, under the Family Planning Accelerator project. SSLE between the two countries started in January 2020 and is still continuing. Both countries started implementation of the learnings (step 4) at the time of preparing this manuscript (December 2021). An independent consultant from Sri Lanka carried out an evaluation, to inform future SSLEs. The evaluation included a desk review on SSLE and family planning in both countries and key informant interviews with Sri Lanka Ministries Health, WHO CO, external partners. A final evaluation of the outcomes/impact is planned in December 2022. The SSLE resulted in a systematic cross-country transfer of knowledge and implementation of the learnings. Sri Lanka implemented a web-based system for logistics management of family planning commodities and Nepal commenced implementing integrated family planning services in a decentralised environment using a lifecycle approach to improve postpartum family planning uptake. The success of this SSLE is attributed to the rigorous methodology, country-led designing of the learning agenda and process, extensive communication amongst the teams, a focus on outcomes, commitment and leadership by ministries of health in both countries. Learning and technical assistance needs of countries can be met by SSLE if national contexts, availability of resources are considered.

Genetic BACH1 deficiency alters mitochondrial function and increases NLRP3 inflammasome activation in mouse macrophages.

BTB-and-CNC homologue 1 (BACH1), a heme-regulated transcription factor, mediates innate immune responses via its functional role in macrophages. BACH1 has recently been shown to modulate mitochondrial metabolism in cancer cells. In the current study, we utilized a proteomics approach and demonstrate that genetic deletion of BACH1 in mouse macrophages is associated with decreased levels of various mitochondrial proteins, particularly mitochondrial complex I. Bioenergetic studies revealed alterations of mitochondrial energy metabolism in BACH1-/- macrophages with a shift towards increased glycolysis and decreased oxidative phosphorylation. Moreover, these cells exhibited enhanced mitochondrial membrane potential and generation of mitochondrial reactive oxygen species (mtROS) along with lower levels of mitophagy. Notably, a higher inducibility of NLRP3 inflammasome activation in response to ATP and nigericin following challenge with lipopolysaccharide (LPS) was observed in BACH1-deficient macrophages compared to wild-type cells. Mechanistically, pharmacological inhibition of mtROS markedly attenuated inflammasome activation. In addition, it is shown that inducible nitric oxide synthase and cyclooxygenase-2, both of which are markedly induced by LPS in macrophages, are directly implicated in BACH1-dependent regulation of NLRP3 inflammasome activation. Taken together, the current findings indicate that BACH1 is critical for immunomodulation of macrophages and may serve as a target for therapeutic approaches in inflammatory disorders.

Prevalence assessment of sexually transmitted infections among pregnant women visiting an antenatal care center of Nepal: Pilot of the World Health Organization's standard protocol for conducting STI prevalence surveys among pregnant women.

INTRODUCTION: Sexually transmitted infections (STIs) are common during pregnancy and can result in adverse delivery and birth outcomes. The purpose of this study was to estimate the prevalence of STIs; Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG), Treponema pallidum (syphilis), Trichomonas vaginalis (trichomoniasis), and Human Immunodeficiency Virus (HIV) among pregnant women visiting an antenatal care center in Nepal. MATERIALS AND METHODS: We adapted and piloted the WHO standard protocol for conducting a prevalence survey of STIs among pregnant women visiting antenatal care center of Dhulikhel Hospital, Nepal. Patient recruitment, data collection, and specimen testing took place between November 2019-March 2020. First catch urine sample was collected from each eligible woman. GeneXpert platform was used for CT and NG testing. Wet-mount microscopy of urine sample was used for detection of trichomoniasis. Serological test for HIV was done by rapid and enzyme-linked immunosorbent assay tests. Serological test for syphilis was done using "nonspecific non-treponemal" and "specific treponemal" antibody tests. Tests for CT, NG and trichomoniasis were done as part of the prevalence study while tests for syphilis and HIV were done as part of the routine antenatal testing. RESULTS: 672 women were approached to participate in the study, out of which 591 (87.9%) met the eligibility criteria and consented to participate. The overall prevalence of any STIs was 8.6% (51/591, 95% CI: 6.3-10.8); 1.5% (95% CI: 0.5-2.5) for CT and 7.1% (95% CI: 5.0-9.2) for trichomoniasis infection. None of the samples tested positive for NG, HIV or syphilis. Prevalence of any STI was not significantly different among women, age ≤ 24 years (10%, 25/229) compared to women age ≥25 years (7.1%, 26/362) (p = 0.08). CONCLUSIONS: The prevalence of trichomoniasis among pregnant women in this sub-urban population of Nepal was high compared to few cases of CT and no cases of NG, syphilis, and HIV. The WHO standard protocol provided a valuable framework for conducting STI surveillance that can be adapted for other countries and populations.

Probiotic properties of lactic acid bacteria isolated from traditionally prepared dry starters of the Eastern Himalayas.

The Himalayan people prepare dry and oval to round-shaped starter cultures to ferment cereals into mild-alcoholic beverages, which contain lactic acid bacteria (LAB) as one of the essential microbiota. There is no report on probiotic characters of LAB isolated from dry starters. Hence, we screened the probiotic and some functional properties of 37 LAB strains isolated from dry starters of the Eastern Himalayas viz. marcha, phab, paa, pee and phut. About 38% of the LAB strains showed high survival rate (> 50%) at pH 3 and 0.3% bile salts. Enterococcus durans BPB21 and SMB7 showed the highest hydrophobicity percentage of 98%. E. durans DMB4 and SMB7 showed maximum cholesterol assimilation activity. About 65% of the LAB strains showed the ability to produce ß galactosidase. Majority of the strains showed phytase activity, whereas none of the strain showed amylase activity. About 86% of LAB strains showed an optimum tolerance of 10% ethanol concentration. Genetic screening of some probiotic and functional marker genes have also been analysed. The occurrence of clp L gene, agu A gene (survival of gastrointestinal tract conditions), apf, mub1 and map A gene (adhesion genes) was higher compared to other genes. The occurrence of bsh gene (bile salt tolerance) was detected in Pediococcus pentosaceus SMB13-1 and Enterococcus faecium BPB11. Gene ped B for pediocin with amplicon size of 375 bp was detected in E. durans DMB13 and Pediococcus acidilactici AKB3. Detection of nutritional marker gene rib A and fol P in some strains showed the potential ability to synthesize riboflavin and folic acid. LAB with probiotic and functional properties may be explored for food industry in future.

Interplay of Heme with Macrophages in Homeostasis and Inflammation.

Macrophages are an integral part of the mononuclear phagocyte system that is critical for maintaining immune homeostasis. They play a key role for initiation and modulation of immunological responses in inflammation and infection. Moreover, macrophages exhibit a wide spectrum of tissue-specific phenotypes in steady-state and pathophysiological conditions. Recent clinical and experimental evidence indicates that the ubiquitous compound heme is a crucial regulator of these cells, e.g., in the differentiation of monocytes to tissue-resident macrophages and/ or in activation by inflammatory stimuli. Notably, heme, an iron containing tetrapyrrole, is essential as a prosthetic group of hemoproteins (e.g., hemoglobin and cytochromes), whereas non-protein bound free or labile heme can be harmful via pro-oxidant, pro-inflammatory, and cytotoxic effects. In this review, it will be discussed how the complex interplay of heme with macrophages regulates homeostasis and inflammation via modulating macrophage inflammatory characteristics and/ or hematopoiesis. A particular focus will be the distinct roles of intra- and extracellular labile heme and the regulation of its availability by heme-binding proteins. Finally, it will be addressed how heme modulates macrophage functions via specific transcriptional factors, in particular the nuclear repressor BTB and CNC homologue (BACH)1 and Spi-C.

Predicting Survival for Patients With Metastatic Disease.

PURPOSE: This prospective study aimed to determine the accuracy of radiation oncologists in predicting the survival of patients with metastatic disease receiving radiation therapy and to understand factors associated with their accuracy. METHODS AND MATERIALS: This single-institution study surveyed 22 attending radiation oncologists to estimate patient survival. Survival predictions were defined as accurate if the observed survival (OS) was within the correct survival prediction category (0-6 months, >6-12 months, >12-24 months, and >24 months). The physicians made survival estimates for each course of radiation, yielding 877 analyzable predictions for 689 unique patients. Data analysis included Stuart's Tau C, logistic regression models, ordinal logistic regression models, and stepwise selection to examine variable interactions. RESULTS: Of the 877 radiation oncologists' predictions, 39.7% were accurate, 26.5% were underestimations, and 33.9% were overestimations. Stuart's Tau C showed low correlation between OS and survival estimates (0.3499), consistent with the inaccuracy reported in the literature. However, results showed less systematic overprediction than reported in the literature. Karnofsky performance status was the most significant predictor of accuracy, with greater accuracy for patients with shorter OS. Estimates were also more accurate for patients with lower Karnofsky performance status. Accuracy by patient age varied by primary site and race. Physician years of experience did not correlate with accuracy. CONCLUSIONS: The sampled radiation oncologists have a 40% accuracy in predicting patient survival. Future investigation should explore how survival estimates influence treatment decisions and how to improve survival prediction accuracy.

Phenotypic and Genotypic Identification of Bacteria Isolated From Traditionally Prepared Dry Starters of the Eastern Himalayas.

Preparation of dry starters for alcohol production is an age-old traditional technology in the Eastern Himalayan regions of east Nepal, the Darjeeling hills, Sikkim, and Arunachal Pradesh in India, and Bhutan. We studied the bacterial diversity in 35 samples of traditionally prepared dry starters, represented by marcha of Nepal, Sikkim, the Darjeeling hills, and Bhutan, phab of Bhutan, and paa, pee, and phut of Arunachal Pradesh, respectively. Populations of bacteria in these starters were 105 to 108 cfu/g. A total of 201 bacterial strains were isolated from starter samples, phenotypically characterized, and their identities confirmed by the 16S rRNA sanger sequencing method. The dominant phylum was Firmicutes (85%), followed by Proteobacteria (9%), and Actinobacteria (6%). Lactic acid bacteria (LAB) (59%) formed the most abundant group, followed by non-LAB (32%) and Gram-negative bacteria (9%). Based on the 16S rRNA gene sequencing result, we identified LAB: Enterococcus durans, E. faecium, E. fecalis, E. hirae, E. lactis, Pediococcus acidilactici, P. pentosaceus, Lactobacillus plantarum subsp. plantarum, Lb. pentosus, Leuconostoc mesenteroides, and Weissella cibaria; non-LAB: Bacillus subtilis subsp. inaquosorum, B. circulans, B. albus, B. cereus, B. nakamurai, B. nitratireducens, B. pseudomycoides, B. zhangzhouensis, Kocuria rosea, Staphylococcus hominis subsp. hominis, S. warneri, S. gallinarum, S. sciuri, Lysinibacillus boronitolerans, Brevibacterium frigoritolerans, and Micrococcus yunnanensis; Gram-negative bacteria: Pseudomonas putida, Klebsiella pneumoniae, Enterobacter hormaechei subsp. xiangfangensis, E. hormaechei subsp. steigerwaltii, and Stenotrophomonas maltophilia. We characterized diversity indexes of the bacterial community present in traditionally prepared dry starters. This is the first report on the bacterial diversity of traditionally dry starters of the Eastern Himalayas by sanger sequencing.

Reduced cognitive deficits after FLASH irradiation of whole mouse brain are associated with less hippocampal dendritic spine loss and neuroinflammation.

AIM: To evaluate the impact of ultra-rapid FLASH mouse whole brain irradiation on hippocampal dendritic spines and neuroinflammation, factors associated with cognitive impairment after brain irradiation. METHODS: We administered 30 Gy whole brain irradiation to C57BL6/J mice in sub-second (FLASH) vs. 240 s conventional delivery time keeping all other parameters constant, using a custom configured clinical linac. Ten weeks post-irradiation, we evaluated spatial and non-spatial object recognition using novel object location and object recognition testing. We measured dendritic spine density by tracing Golgi-stained hippocampal neurons and evaluated neuroinflammation by CD68 immunostaining, a marker of activated microglia, and expression of 10 pro-inflammatory cytokines using a multiplex immunoassay. RESULTS: At ten weeks post-irradiation, compared to unirradiated controls, conventional delivery time irradiation significantly impaired novel object location and recognition tasks whereas the same dose given in FLASH delivery did not. Conventional delivery time, but not FLASH, was associated with significant loss of dendritic spine density in hippocampal apical dendrites, with a similar non-significant trend in basal dendrites. Conventional delivery time was associated with significantly increased CD68-positive microglia compared to controls whereas FLASH was not. Conventional delivery time was associated with significant increases in 5 of 10 pro-inflammatory cytokines in the hippocampus (and non-significant increases in another 3), whereas FLASH was associated with smaller increases in only 3. CONCLUSION: Reduced cognitive impairment and associated neurodegeneration were observed with FLASH compared to conventional delivery time irradiation, potentially through decreased induction of neuroinflammation, suggesting a promising approach to increasing therapeutic index in radiation therapy of brain tumors.

Human and murine macrophages exhibit differential metabolic responses to lipopolysaccharide - A divergent role for glycolysis.

Macrophages adopt different phenotypes in response to microenvironmental changes, which can be principally classified into inflammatory and anti-inflammatory states. Inflammatory activation of macrophages has been linked with metabolic reprogramming from oxidative phosphorylation to aerobic glycolysis. In contrast to mouse macrophages, little information is available on the link between metabolism and inflammation in human macrophages. In the current report it is demonstrated that lipopolysaccharide (LPS)-activated human peripheral blood monocyte-derived macrophages (hMDMs) fail to undergo metabolic reprogramming towards glycolysis, but rely on oxidative phosphorylation for the generation of ATP. By contrast, activation by LPS led to an increased extracellular acidification rate (glycolysis) and decreased oxygen consumption rate (oxidative phosphorylation) in mouse bone marrow-derived macrophages (mBMDMs). Mitochondrial bioenergetics after LPS stimulation in human macrophages was unchanged, but was markedly impaired in mouse macrophages. Furthermore, treatment with 2-deoxyglucose, an inhibitor of glycolysis, led to cell death in mouse, but not in human macrophages. Finally, glycolysis appeared to be critical for LPS-mediated induction of the anti-inflammatory cytokine interleukin-10 in both human and mouse macrophages. In summary, these findings indicate that LPS-induced immunometabolism in human macrophages is different to that observed in mouse macrophages.

Automated Survival Prediction in Metastatic Cancer Patients Using High-Dimensional Electronic Medical Record Data.

BACKGROUND: Oncologists use patients' life expectancy to guide decisions and may benefit from a tool that accurately predicts prognosis. Existing prognostic models generally use only a few predictor variables. We used an electronic medical record dataset to train a prognostic model for patients with metastatic cancer. METHODS: The model was trained and tested using 12 588 patients treated for metastatic cancer in the Stanford Health Care system from 2008 to 2017. Data sources included provider note text, labs, vital signs, procedures, medication orders, and diagnosis codes. Patients were divided randomly into a training set used to fit the model coefficients and a test set used to evaluate model performance (80%/20% split). A regularized Cox model with 4126 predictor variables was used. A landmarking approach was used due to the multiple observations per patient, with t0 set to the time of metastatic cancer diagnosis. Performance was also evaluated using 399 palliative radiation courses in test set patients. RESULTS: The C-index for overall survival was 0.786 in the test set (averaged across landmark times). For palliative radiation courses, the C-index was 0.745 (95% confidence interval [CI] = 0.715 to 0.775) compared with 0.635 (95% CI = 0.601 to 0.669) for a published model using performance status, primary tumor site, and treated site (two-sided P < .001). Our model's predictions were well-calibrated. CONCLUSIONS: The model showed high predictive performance, which will need to be validated using external data. Because it is fully automated, the model can be used to examine providers' practice patterns and could be deployed in a decision support tool to help improve quality of care.

Labile Heme Aggravates Renal Inflammation and Complement Activation After Ischemia Reperfusion Injury.

Background: Ischemia reperfusion injury (IRI) plays a major role in solid organ transplantation. The length of warm ischemia time is critical for the extent of tissue damage in renal IRI. In this experimental study we hypothesized that local release of labile heme in renal tissue is triggered by the duration of warm ischemia (15 vs. 45 min IRI) and mediates complement activation, cytokine release, and inflammation. Methods: To induce IRI, renal pedicle clamping was performed in male C57BL/6 mice for short (15 min) or prolonged (45 min) time periods. Two and 24 h after experimental ischemia tissue injury labile heme levels in the kidney were determined with an apo-horseradish peroxidase assay. Moreover, renal injury, cytokines, and C5a and C3a receptor (C5aR, C3aR) expression were determined by histology, immunohistochemistry and qPCR, respectively. In addition, in vitro studies stimulating bone marrow-derived macrophages with LPS and the combination of LPS and heme were performed and cytokine expression was measured. Results: Inflammation and local tissue injury correlated with the duration of warm ischemia time. Labile heme concentrations in renal tissue were significantly higher after prolonged (45 min) as compared to short (15 min) IRI. Notably, expression of the inducible heme-degrading enzyme heme oxygenase-1 (HO-1) was up-regulated in kidneys after prolonged, but not after short IRI. C5aR, the pro-inflammatory cytokines IL-6 and TNF-α as well as pERK were up-regulated after prolonged, but not after short ischemia times. Consecutively, neutrophil infiltration and up-regulation of pro-fibrotic cytokines such as CTGF and PAI were more pronounced in prolonged IRI in comparison to short IRI. In vitro stimulation of macrophages with LPS revealed that IL-6 expression was enhanced in the presence of heme. Finally, administration of the heme scavenger human serum albumin (HSA) reduced the expression of pro-inflammatory cytokines, C3a receptor and improved tubular function indicated by enhanced alpha 1 microglobulin (A1M) absorption after IRI. Conclusions: Our data show that prolonged duration of warm ischemia time increased labile heme levels in the kidney, which correlates with IRI-dependent inflammation and up-regulation of anaphylatoxin receptor expression.

The role of bone marrow and spleen irradiation in the development of acute hematologic toxicity during chemoradiation for esophageal cancer.

PURPOSE: The purpose of this study was to determine the impact of splenic and thoracic bone marrow irradiation on hematologic toxicity in the setting of chemoradiation therapy for esophageal cancer. METHODS AND MATERIALS: We analyzed 60 patients with carcinoma of the distal esophagus or gastroesophageal junction who received concurrent chemoradiation in the preoperative or definitive setting. Dosimetric and volumetric parameters were calculated for the spleen, thoracic spine, and posterior ribs. The primary endpoint was grade ≥3 hematologic toxicity (HT3+). Associations were assessed using logistic and linear regression models. RESULTS: Twenty-one patients (35%) experienced HT3+, including 18 patients with leukopenia and 5 with thrombocytopenia. Higher spleen V5-V20 was correlated with a lower risk of HT3+ on multivariable analysis (odds ratio: 0.83 per 10 cm3 increase in V10; P = .013). A dose-dependent decrease in spleen volume was observed after radiation therapy, and a greater decrease was independently associated with a lower risk of HT3+ (odds ratio: 0.93 per 1% volume decrease; P = .014). Dosimetric parameters of the thoracic spine were not significantly associated with HT3+. CONCLUSIONS: A greater decrease in spleen size after radiation therapy and a higher spleen V5-V20 were independently associated with a lower risk of severe hematologic toxicity. Splenic irradiation may mitigate leukopenia associated with chemoradiation therapy.

Enhanced activation of interleukin-10, heme oxygenase-1, and AKT in C5aR2-deficient mice is associated with protection from ischemia reperfusion injury-induced inflammation and fibrosis.

Severe ischemia reperfusion injury (IRI) results in rapid complement activation, acute kidney injury and progressive renal fibrosis. Little is known about the roles of the C5aR1 and C5aR2 complement receptors in IRI. In this study C5aR1-/- and C5aR2-/- mice were compared to the wild type in a renal IRI model leading to renal fibrosis. C5a receptor expression, kidney morphology, inflammation, and fibrosis were measured in different mouse strains one, seven and 21 days after IRI. Renal perfusion was evaluated by functional magnetic resonance imaging. Protein abundance and phosphorylation were assessed with high content antibody microarrays and Western blotting. C5aR1 and C5aR2 were increased in damaged tubuli and even more in infiltrating leukocytes after IRI in kidneys of wild-type mice. C5aR1-/- and C5aR2-/- animals developed less IRI-induced inflammation and showed better renal perfusion than wild-type mice following IRI. C5aR2-/- mice, in particular, had enhanced tubular and capillary regeneration with less renal fibrosis. Anti-inflammatory IL-10 and the survival/growth kinase AKT levels were especially high in kidneys of C5aR2-/- mice following IRI. LPS caused bone marrow-derived macrophages from C5aR2-/- mice to release IL-10 and to express the stress response enzyme heme oxygenase-1. Thus, C5aR1 and C5aR2 have overlapping actions in which the kidneys of C5aR2-/- mice regenerate better than those in C5aR1-/- mice following IRI. This is mediated, at least in part, by differential production of IL-10, heme oxygenase-1 and AKT.

Analysis of bacterial and fungal communities in Marcha and Thiat, traditionally prepared amylolytic starters of India.

Marcha and thiat are traditionally prepared amylolytic starters use for production of various ethnic alcoholic beverages in Sikkim and Meghalaya states in India. In the present study we have tried to investigate the bacterial and fungal community composition of marcha and thiat by using high throughput sequencing. Characterization of bacterial community depicts phylum Proteobacteria is the most dominant in both marcha (91.4%) and thiat (53.8%), followed by Firmicutes, and Actinobacteria. Estimates of fungal community composition showed Ascomycota as the dominant phylum. Presence of Zygomycota in marcha distinguishes it from the thiat. The results of NGS analysis revealed dominance of yeasts in marcha whereas molds out numbers in case of thiat. This is the first report on microbial communities of traditionally prepared amylolytic starters of India using high throughput sequencing.

Molecular Contrast-Enhanced Ultrasound Imaging of Radiation-Induced P-Selectin Expression in Healthy Mice Colon.

PURPOSE: To evaluate the feasibility of using molecular contrast-enhanced ultrasound (mCEUS) to image radiation (XRT)-induced expression of cell adhesion molecules that mediate inflammatory response to XRT in healthy mouse colon tissue. METHODS AND MATERIALS: The colons of male BALB/c mice (aged 6-8 weeks, n=9) were irradiated with 14 Gy using a Kimtron IC-225 x-ray irradiator operating at 225 kV/13.0 mA at a dose rate of 0.985 Gy/min. The head and thorax regions were shielded during irradiation. A second control cohort of mice was left untreated (n=6). Molecular CEUS was carried out before and 24 hours after irradiation using a VEVO2100 system and MS250 21-MHz center frequency transducer. Each imaging session comprised mCEUS imaging with P-selectin targeted microbubbles and control microbubbles targeted with an isotype control IgG. Quantification of mCEUS was carried out by measuring the differential targeted enhancement (dTE) parameter. The perfusion parameters peak enhancement and area under the curve were also extracted from the initial injection bolus. Animals were sacrificed at 24 hours and the colon was resected for immunohistochemistry analysis (P-selectin/CD31-stained vessel). RESULTS: For P-selectin targeted microbubble, a significant increase (40 a.u.; P=.013) in dTE (P-dTE) was observed in irradiated mice over 24 hours. In contrast, a nonsignificant change in P-selectin dTE was observed in control mice. For control microbubbles, no significant difference in the IgG dTE parameter was noted in treated and control animals over 24 hours. A nonsignificant increase in the peak enhancement and area under the curve perfusion parameters associated with blood volume was noted in animals treated with radiation. Quantitative histology indicated significantly elevated P-selectin expression per blood vessel (36% in treated; 14% in control). CONCLUSION: Our results confirm the feasibility of using mCEUS for imaging of XRT-induced expression of P-selectin as a potential approach to monitoring healthy tissue inflammatory damage during radiation therapy.

Ocular morbidity among children studying in private schools of Kathmandu valley: A prospective cross sectional study.

A prospective cross sectional study of students of two private schools was done and students were screened in detail. A total of 1816 students aged 5 to 16 years were evaluated, out of which 52.8% were males and 47.2 % were females. Among the total, 65.8% had no ocular abnormalities and 34.2% had some form of ocular disorders. Refractive error was the commonest problem seen accounting for 21.9% out of total, followed by infective disorders, which accounted for 7.2%, 3.5% of them were noted to have Orthoptic problem including various types of strabismus, 2.2% were color blind, 2.6% were found to have various other disorders. The prevalence of refractive error among private school children seems to be higher. Color blindness also seems to be prevalent among these children. This kind of school screening would help in detecting the eye problems timely and thus would reduce the ocular morbidity as well as prevent children from going blind unnecessarily.