Activin A-Expressing Polymorphonuclear Myeloid-Derived Suppressor Cells Infiltrate Skeletal and Cardiac Muscle and Promote Cancer Cachexia.

Cachexia is a major cause of death in cancer and leads to wasting of cardiac and skeletal muscle, as well as adipose tissue. Various cellular and soluble mediators have been postulated in driving cachexia; however, the specific mechanisms behind this muscle wasting remain poorly understood. In this study, we found polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) to be critical for the development of cancer-associated cachexia. Significant expansion of PMN-MDSCs was observed in the cardiac and skeletal muscles of cachectic murine models. Importantly, the depletion of this cell subset, using depleting anti-Ly6G Abs, attenuated this cachectic phenotype. To elucidate the mechanistic involvement of PMN-MDSCs in cachexia, we examined major mediators, that is, IL-6, TNF-α, and arginase 1. By employing a PMN-MDSC-specific Cre-recombinase mouse model, we showed that PMN-MDSCs were not maintained by IL-6 signaling. In addition, PMN-MDSC-mediated cardiac and skeletal muscle loss was not abrogated by deficiency in TNF-α or arginase 1. Alternatively, we found PMN-MDSCs to be critical producers of activin A in cachexia, which was noticeably elevated in cachectic murine serum. Moreover, inhibition of the activin A signaling pathway completely protected against cardiac and skeletal muscle loss. Collectively, we demonstrate that PMN-MDSCs are active producers of activin A, which in turn induces cachectic muscle loss. Targeting this immune/hormonal axis will allow the development of novel therapeutic interventions for patients afflicted with this debilitating syndrome.

Butyrate driven raft disruption trots off enteric pathogen invasion: possible mechanism of colonization resistance.

The gut microbiome derived short chain fatty acids perform multitude of functions to maintain gut homeostasis. Here we studied how butyrate stymie enteric bacterial invasion in cell using a simplistic binary model. The surface of the mammalian cells is enriched with microdomains rich in cholesterol that are known as rafts and act as entry points for pathogens. We showed that sodium butyrate treated RAW264.7 cells displayed reduced membrane cholesterol and less cholera-toxin B binding coupled with increased membrane fluidity compared to untreated cells indicating that reduced membrane cholesterol caused disruption of lipid rafts. The implication of such cellular biophysical changes on the invasion of enteric pathogenic bacteria was assessed. Our study showed, in comparison to untreated cells, butyrate-treated cells significantly reduced the invasion of Shigella and Salmonella, and these effects were found to be reversed by liposomal cholesterol treatment, increasing the likelihood that the rafts' function against bacterial invasion. The credence of ex vivo studies found to be in concordance in butyrate fed mouse model as evident from the significant drift towards a protective phenotype against virulent enteric pathogen invasion as compared to untreated mice. To produce a cytokine balance towards anti-inflammation, butyrate-treated mice produced more of the gut tissue anti-inflammatory cytokine IL-10 and less of the pro-inflammatory cytokines TNF-α, IL-6, and IFN-γ. In histological studies of Shigella infected gut revealed a startling observation where number of neutrophils infiltration was noted which was correlated with the pathology and was essentially reversed by butyrate treatment. Our results ratchet up a new dimension of our understanding how butyrate imparts resistance to pathogen invasion in the gut.

Prenatal arsenic exposure stymies gut butyrate production and enhances gut permeability in post natal life even in absence of arsenic deftly through miR122-Occludin pathway.

This discourse attempts to capture a few important dimensions of gut physiology like microbial homeostasis, short chain fatty acid (SCFA) production, occludin expression, and gut permeability in post-natal life of mice those received arsenic only during pre-natal life. Adult Balb/c mice were fed with 4 ppm arsenic trioxide in drinking water during breeding and gestation. After the birth of the pups, the arsenic water was withdrawn and replaced with clean drinking water. The pups were allowed to grow for 28 days (pAs-mice) and age matched Balb/c mice which were never exposed to arsenic served as control The pAs-mice showed a striking reduction in Firmicutes to Bacteroidetes (F/B) ratio coupled with a decrease in tight junction protein, occludin resulting in an increase in gut permeability, increased infiltration of inflammatory cells in the colon and decrease in common SCFAs in which butyrate reduction was quite prominent in fecal samples as compared to normal control. The above phenotypes of pAs-mice were mostly reversed by supplementing 5% sodium butyrate (w/w) with food from 21st to 28th day. The ability of butyrate in enhancing occludin expression, in particular, was dissected further. As miR122 causes degradation of Occludin mRNA, we transiently overexpressed miR122 by injecting appropriate plasmids and showed reversal of butyrate effects in pAs-mice. Thus, pre-natal arsenic exposure orchestrates variety of effects by decreasing butyrate in pAs-mice leading to increased permeability due to reduced occludin expression. Our research adds a new dimension to our understanding that pre-natal arsenic exposure imprints in post-natal life while there was no further arsenic exposure.

The NFκB signaling system in the generation of B-cell subsets: from germinal center B cells to memory B cells and plasma cells.

Memory B cells and antibody-secreting cells are the two prime effector B cell populations that drive infection- and vaccine-induced long-term antibody-mediated immunity. The antibody-mediated immunity mostly relies on the formation of specialized structures within secondary lymphoid organs, called germinal centers (GCs), that facilitate the interactions between B cells, T cells, and antigen-presenting cells. Antigen-activated B cells may proliferate and differentiate into GC-independent plasmablasts and memory B cells or differentiate into GC B cells. The GC B cells undergo proliferation coupled to somatic hypermutation of their immunoglobulin genes for antibody affinity maturation. Subsequently, affinity mature GC B cells differentiate into GC-dependent plasma cells and memory B cells. Here, we review how the NFκB signaling system controls B cell proliferation and the generation of GC B cells, plasmablasts/plasma cells, and memory B cells. We also identify and discuss some important unanswered questions in this connection.

B Cells Promote T Cell Immunosenescence and Mammalian Aging Parameters.

A dysregulated adaptive immune system is a key feature of aging, and is associated with age-related chronic diseases and mortality. Most notably, aging is linked to a loss in the diversity of the T cell repertoire and expansion of activated inflammatory age-related T cell subsets, though the main drivers of these processes are largely unknown. Here, we find that T cell aging is directly influenced by B cells. Using multiple models of B cell manipulation and single-cell omics, we find B cells to be a major cell type that is largely responsible for the age-related reduction of naive T cells, their associated differentiation towards pathogenic immunosenescent T cell subsets, and for the clonal restriction of their T cell receptor (TCR). Accordingly, we find that these pathogenic shifts can be therapeutically targeted via CD20 monoclonal antibody treatment. Mechanistically, we uncover a new role for insulin receptor signaling in influencing age-related B cell pathogenicity that in turn induces T cell dysfunction and a decline in healthspan parameters. These results establish B cells as a pivotal force contributing to age-associated adaptive immune dysfunction and healthspan outcomes, and suggest new modalities to manage aging and related multi-morbidity.

PDMS hydrogel-coated tissue culture plates for studying the impact of substrate stiffness on dendritic cell function.

The mechanical properties of polydimethylsiloxane hydrogels can be tuned to mimic physiological tensions, an underappreciated environmental parameter in immunology studies. We describe a workflow to prepare PDMS-coated tissue culture plates with biologically relevant substrate stiffness, and the use of these hydrogel plates to condition isolated primary splenic CD11c+ dendritic cells (DC). Finally, we suggest downstream applications to study the impact of substrate stiffness on DC function and metabolism. The protocol could be adapted to study other mechanosensitive immune cell subsets. For complete details on the use and execution of this protocol, please refer to Chakraborty et al. (2021).

AUF-1 knockdown in mice undermines gut microbial butyrate-driven hypocholesterolemia through AUF-1-Dicer-1-mir-122 hierarchy.

Introduction and objective: Cholesterol homeostasis is a culmination of cellular synthesis, efflux, and catabolism to important physiological entities where short chain fatty acid, butyrate embodied as a key player. This discourse probes the mechanistic molecular details of butyrate action in maintaining host-cholesterol balance. Methods: Hepatic mir-122 being the most indispensable regulator of cholesterol metabolic enzymes, we studied upstream players of mir-122 biogenesis in the presence and absence of butyrate in Huh7 cells and mice model. We synthesized unique self-transfecting GMO (guanidinium-morpholino-oligo) linked PMO (Phosphorodiamidate-Morpholino Oligo)-based antisense cell-penetrating reagent to selectively knock down the key player in butyrate mediated cholesterol regulation. Results: We showed that butyrate treatment caused upregulation of RNA-binding protein, AUF1 resulting in RNase-III nuclease, Dicer1 instability, and significant diminution of mir-122. We proved the importance of AUF1 and sequential downstream players in AUF1-knock-down mice. Injection of GMO-PMO of AUF1 in mouse caused near absence of AUF1 coupled with increased Dicer1 and mir-122, and reduced serum cholesterol regardless of butyrate treatment indicating that butyrate acts through AUF1. Conclusion: The roster of intracellular players was as follows: AUF1-Dicer1-mir-122 for triggering butyrate driven hypocholesterolemia. To our knowledge this is the first report linking AUF-1 with cholesterol biogenesis.

Corona-Nidaan: lightweight deep convolutional neural network for chest X-Ray based COVID-19 infection detection.

The coronavirus COVID-19 pandemic is today's major public health crisis, we have faced since the Second World War. The pandemic is spreading around the globe like a wave, and according to the World Health Organization's recent report, the number of confirmed cases and deaths are rising rapidly. COVID-19 pandemic has created severe social, economic, and political crises, which in turn will leave long-lasting scars. One of the countermeasures against controlling coronavirus outbreak is specific, accurate, reliable, and rapid detection technique to identify infected patients. The availability and affordability of RT-PCR kits remains a major bottleneck in many countries, while handling COVID-19 outbreak effectively. Recent findings indicate that chest radiography anomalies can characterize patients with COVID-19 infection. In this study, Corona-Nidaan, a lightweight deep convolutional neural network (DCNN), is proposed to detect COVID-19, Pneumonia, and Normal cases from chest X-ray image analysis; without any human intervention. We introduce a simple minority class oversampling method for dealing with imbalanced dataset problem. The impact of transfer learning with pre-trained CNNs on chest X-ray based COVID-19 infection detection is also investigated. Experimental analysis shows that Corona-Nidaan model outperforms prior works and other pre-trained CNN based models. The model achieved 95% accuracy for three-class classification with 94% precision and recall for COVID-19 cases. While studying the performance of various pre-trained models, it is also found that VGG19 outperforms other pre-trained CNN models by achieving 93% accuracy with 87% recall and 93% precision for COVID-19 infection detection. The model is evaluated by screening the COVID-19 infected Indian Patient chest X-ray dataset with good accuracy.

Mechanical Stiffness Controls Dendritic Cell Metabolism and Function.

Stiffness in the tissue microenvironment changes in most diseases and immunological conditions, but its direct influence on the immune system is poorly understood. Here, we show that static tension impacts immune cell function, maturation, and metabolism. Bone-marrow-derived and/or splenic dendritic cells (DCs) grown in vitro at physiological resting stiffness have reduced proliferation, activation, and cytokine production compared with cells grown under higher stiffness, mimicking fibro-inflammatory disease. Consistently, DCs grown under higher stiffness show increased activation and flux of major glucose metabolic pathways. In DC models of autoimmune diabetes and tumor immunotherapy, tension primes DCs to elicit an adaptive immune response. Mechanistic workup identifies the Hippo-signaling molecule, TAZ, as well as Ca2+-related ion channels, including potentially PIEZO1, as important effectors impacting DC metabolism and function under tension. Tension also directs the phenotypes of monocyte-derived DCs in humans. Thus, mechanical stiffness is a critical environmental cue of DCs and innate immunity.

Caring for cancer patients in the Covid pandemic: choosing between the devil and deep sea.

BACKGROUND: Healthcare is an essential service at any time more so in the crisis like Covid. With increase in number of cases and mortality from Covid, the primary focus is shifted to the management of the Covid crisis and other health emergencies thus affecting normal health services and routine treatment of other diseases like cancer. METHODS: This article reviews the published literature and guidelines on Covid and cancer and discusses them to optimize the care of cancer patients during Covid pandemic to improve treatment outcomes. RESULTS: The results of the review of published literature show a twofold increase in probability of getting CoV2 infection by the cancer patients and a four-fold increase in chance of death. On the other hand, if left untreated a 20% increase in cancer death is expected. Data further show that none of the medicines like remdesivir, hydroxy chloroquin, dexamethasone, or azithromycin improves survival and response to Covid in cancer patients. Surgical results too show similar outcome before and after the pandemic though most of these report on highly selected patients populations. CONCLUSIONS: The Covid 2019 pandemic places cancer patients in a very difficult situation wherein if they seek treatment, they are exposing themselves to a risk of developing CoV2 infection and if they do not, the probability of dying without treatment increases. Hence, for them it is a choice between the devil and deep sea, and it is for the healthcare providers to triage patients and treat who cannot wait even though the data from the carefully selected cohort of patients show no increase in mortality or morbidity from treatment during Covid.

Prenatal arsenic exposure interferes in postnatal immunocompetence despite an absence of ongoing arsenic exposure.

Arsenic (As) readily crosses the placenta and exposure of the fetus may cause adverse consequences later in life, including immunomodulation. In the current study, the question was asked how the immune repertoire might respond in postnatal life when there is no further As exposure. Here, pregnant mice (Balb/c [H-2d]) were exposed to arsenic trioxide (As2O3) through their drinking water from time of conception until parturition. Their offspring, 4-week-old mice who had not been exposed again to As, were used for functional analyses of innate, humoral and cellular immunity. Compared to cells from non-As-exposed dam offspring, isolated peritoneal macro-phages (Mϕ) displayed no differences in T-cell stimulating ability. Levels of circulating IgG2a but not IgG1 were decreased in As-exposed dam offspring as compared to control offspring counterparts. Mixed-leukocyte reactions (MLR) indicated that CD4+ T-cells from the prenatal As-exposed mice were significantly less responsive to allogenic stimulation as evidenced by decreases in interferon (IFN)-γ and IL-2 production and in expression of CD44 and CD69 (but not CD25) activation markers. Interestingly, the Mϕ from the prenatal As-exposed mice were capable of stimulating normal allogenic T-cells, indicating that T-cells from these mice were refractory to allogenic signals. There was also a significant decrease in absolute numbers of splenic CD4+ and CD8+ T-cells due to prenatal As exposure (as compared to control). Lastly, the impaired immune function of the prenatal As-exposed mice was correlated with a very strong susceptibility to Escherichia coli infection. Taken together, the data from this study clearly show that in utero As exposure may continue to perpetuate a dampening effect on the immune repertoire of offspring, even into the early stages of postnatal life.

Gut-associated IgA+ immune cells regulate obesity-related insulin resistance.

The intestinal immune system is emerging as an important contributor to obesity-related insulin resistance, but the role of intestinal B cells in this context is unclear. Here, we show that high fat diet (HFD) feeding alters intestinal IgA+ immune cells and that IgA is a critical immune regulator of glucose homeostasis. Obese mice have fewer IgA+ immune cells and less secretory IgA and IgA-promoting immune mediators. HFD-fed IgA-deficient mice have dysfunctional glucose metabolism, a phenotype that can be recapitulated by adoptive transfer of intestinal-associated pan-B cells. Mechanistically, IgA is a crucial link that controls intestinal and adipose tissue inflammation, intestinal permeability, microbial encroachment and the composition of the intestinal microbiome during HFD. Current glucose-lowering therapies, including metformin, affect intestinal-related IgA+ B cell populations in mice, while bariatric surgery regimen alters the level of fecal secretory IgA in humans. These findings identify intestinal IgA+ immune cells as mucosal mediators of whole-body glucose regulation in diet-induced metabolic disease.

A bronchoprotective role for Rgs2 in a murine model of lipopolysaccharide-induced airways inflammation.

BACKGROUND: Asthma exacerbations are associated with the recruitment of neutrophils to the lungs. These cells release proteases and mediators, many of which act at G protein-coupled receptors (GPCRs) that couple via Gq to promote bronchoconstriction and inflammation. Common asthma therapeutics up-regulate expression of the regulator of G protein signalling (RGS), RGS2. As RGS2 reduces signaling from Gq-coupled GPCRs, we have defined role(s) for this GTPase-activating protein in an acute neutrophilic model of lung inflammation. METHODS: Wild type and Rgs2 -/- C57Bl6 mice were exposed to nebulized lipopolysaccharide (LPS). Lung function (respiratory system resistance and compliance) was measured using a SCIREQ flexivent small animal ventilator. Lung inflammation was assessed by histochemistry, cell counting and by cytokine and chemokine expression in bronchoalveolar lavage (BAL) fluid. RESULTS: Lipopolysaccharide inhalation induced transient airways hyperreactivity (AHR) and neutrophilic lung inflammation. While AHR and inflammation was greatest 3 h post-LPS exposure, BAL neutrophils persisted for 24 h. At 3 h post-LPS inhalation, multiple inflammatory cytokines (CSF2, CSF3, IL6, TNF) and chemokines (CCL3, CCL4, CXCL1, CXCL2) were highly expressed in the BAL fluid, prior to declining by 24 h. Compared to wild type counterparts, Rgs2 -/- mice developed significantly greater airflow resistance in response to inhaled methacholine (MCh) at 3 h post-LPS exposure. At 24 h post-LPS exposure, when lung function was recovering in the wild type animals, MCh-induced resistance was increased, and compliance decreased, in Rgs2 -/- mice. Thus, Rgs2 -/- mice show AHR and stiffer lungs 24 h post-LPS exposure. Histological markers of inflammation, total and differential cell counts, and major cytokine and chemokine expression in BAL fluid were similar between wild type and Rgs2 -/- mice. However, 3 and 24 h post-LPS exposure, IL12B expression was significantly elevated in BAL fluid from Rgs2 -/- mice compared to wild type animals. CONCLUSIONS: While Rgs2 is bronchoprotective in acute neutrophilic inflammation, no clear anti-inflammatory effect was apparent. Nevertheless, elevated IL12B expression in Rgs2 -/- animals raises the possibility that RGS2 could dampen Th1 responses. These findings indicate that up-regulation of RGS2, as occurs in response to inhaled corticosteroids and long-acting ß2-adrenoceptor agonists, may be beneficial in acute neutrophilic exacerbations of airway disease, including asthma.

Serum Ferritin: A Backstage Weapon in Diagnosis of Dengue Fever.

AIMS: This retrospective study evaluates ferritin as a surrogate marker for dengue infection (NS1 and IgM negative stage) as opposed to other febrile illnesses of infective or inflammatory etiology (OFI). METHODOLOGY: Data of all patients admitted to medical ward and medical ITU during the dengue outbreak were collected. Patients admitted between 5 and 10 days of febrile illness without a diagnosis were included. Patients with NS1 positivity (Days 2-8) and/or positive IgM for dengue (Days 6-10) were considered to be dengue cases and those with other confirmed diagnoses were considered in the OFI group. Ferritin, CRP, TC of WBC, platelet count, SGOT, SGPT, and albumin levels were analysed for both groups. RESULTS: We examined 30 cases of clinically and serologically confirmed dengue fever and 22 cases of OFI. Ferritin level in dengue cohort was significantly higher than the OFI group (p < 0.0001). The best cut-off for ferritin level to differentiate dengue from OFI was found to be 1291. The sensitivity at this cut-off is 82.6% and the specificity at this cut-off is 100%. CONCLUSION: Ferritin may serve as a significant marker for differentiating between dengue fever and OFI, in absence of a positive NS1 antigen or a positive IgM antibody for dengue.

Flavonoid Enriched Fraction of Campylandra aurantiaca Attenuates Carbon Tetrachloride Induced Oxidative DNA Damage in Mouse Peritoneal Macrophages in Animal Model.

BACKGROUND: Recent studies have sought to draw attention of biological activity of Campylandra aurantiaca. The aim of the present study was to evaluate the effect of flavonoid enriched fraction of Campylandra aurantiaca (FEFCA) on in vitro and in vivo antioxidant and DNA protective effect in mouse peritoneal macrophages cells. METHODS: FEFCA was characterized by HPLC analysis. The in vitro antioxidant activities of FEFCA was measured by different in vitro assays like 1, 1-diphenyl-2-picrylhydrazil radical (DPPH), superoxide anions, nitric oxide and hydroxyl radicals scavenging methods. Isolated mouse peritoneal macrophages were oxidized by carbon tetra chloride (CCl4) in animal model; subsequently the protective effect of FEFCA was determined in terms of estimation of antioxidant enzyme and the damage to DNA of the cells. RESULTS: FEFCA exhibited both in vitro antioxidant activities in a concentration dependent manner. FEFCA significantly (*p<0.05) attenuated the oxidative DNA damage of mouse peritoneal macrophage cells induced by CCl4 in an in vivo assay. CONCLUSION: Therefore FEFCA showed good free radical scavenging activity as well as reduced oxidative DNA damage in mouse peritoneal macrophages in animal model.

Protective Roles for RGS2 in a Mouse Model of House Dust Mite-Induced Airway Inflammation.

The GTPase-accelerating protein, regulator of G-protein signalling 2 (RGS2) reduces signalling from G-protein-coupled receptors (GPCRs) that signal via Gαq. In humans, RGS2 expression is up-regulated by inhaled corticosteroids (ICSs) and long-acting ß2-adrenoceptor agonists (LABAs) such that synergy is produced in combination. This may contribute to the superior clinical efficacy of ICS/LABA therapy in asthma relative to ICS alone. In a murine model of house dust mite (HDM)-induced airways inflammation, three weeks of intranasal HDM (25 µg, 3×/week) reduced lung function and induced granulocytic airways inflammation. Compared to wild type animals, Rgs2-/- mice showed airways hyperresponsiveness (increased airways resistance and reduced compliance). While HDM increased pulmonary inflammation observed on hematoxylin and eosin-stained sections, there was no difference between wild type and Rgs2-/- animals. HDM-induced mucus hypersecretion was also unaffected by RGS2 deficiency. However, inflammatory cell counts in the bronchoalveolar lavage fluid of Rgs2-/- animals were significantly increased (57%) compared to wild type animals and this correlated with increased granulocyte (neutrophil and eosinophil) numbers. Likewise, cytokine and chemokine (IL4, IL17, IL5, LIF, IL6, CSF3, CXCLl, CXCL10 and CXCL11) release was increased by HDM exposure. Compared to wild type, Rgs2-/- animals showed a trend towards increased expression for many cytokines/chemokines, with CCL3, CCL11, CXCL9 and CXCL10 being significantly enhanced. As RGS2 expression was unaffected by HDM exposure, these data indicate that RGS2 exerts tonic bronchoprotection in HDM-induced airways inflammation. Modest anti-inflammatory and anti-remodelling roles for RGS2 are also suggested. If translatable to humans, therapies that maximize RGS2 expression may prove advantageous.

Amelioration of oxidative DNA damage in mouse peritoneal macrophages by Hippophae salicifolia due to its proton (H(+)) donation capability: Ex vivo and in vivo studies.

INTRODUCTION: The present study evaluates the antioxidant effect of methanol extract of Hippophae salicifolia (MEHS) bark with special emphasis on its role on oxidative DNA damage in mouse peritoneal macrophages. MATERIAL AND METHODS: In vitro antioxidant activity was estimated by standard antioxidant assays whereas the antioxidant activity concluded the H(+) donating capacity. Mouse erythrocytes' hemolysis and peritoneal macrophages' DNA damage were determined spectrophotometrically. In vivo antioxidant activity of MEHS was determined in carbon tetrachloride-induced mice by studying its effect on superoxide anion production in macrophages cells, superoxide dismutase in the cell lysate, DNA damage, lipid peroxidation, and reduces glutathione. RESULTS: The extract showed good in vitro antioxidant activities whereas the inhibitory concentrations values ranged from 5.80 to 106.5 µg/ml. MEHS significantly (P < 0.05) attenuated the oxidative DNA damage. It also attenuated the oxidative conversion of hemoglobin to methemoglobin and elevation of enzymatic and nonenzymatic antioxidant in cells. CONCLUSION: The result indicates MEHS has good in vitro-in vivo antioxidant property as well as the protective effect on DNA and red blood cell may be due to its H(+) donating property.

Regulation of apoptosis through bcl-2/bax proteins expression and DNA damage by Zanthoxylum alatum.

CONTEXT: Many of the major chemotherapeutic agents are secondary metabolites found in nature. Zanthoxylum alatum Roxb. (Rutaceae) is traditionally used in the treatment of various diseases. OBJECTIVE: The present study evaluates the apoptotic activity of methanol extract of Z. alatum (MEZA) on Ehrlich ascites tumor (EAT) in Swiss albino mice. MATERIALS AND METHODS: The presence of flavonoids in MEZA was standardized by HPLC. The in vitro cytotoxicity of MEZA was measured by the MTT assay. The in vivo antitumor activity of MEZA (100 and 200 mg/kg b.w., i.p. for 9 days) was also evaluated. On the 10th day, EAT tumor volume, cell viability, and hematological parameters were assayed. Apoptotic morphology was determined by acredine orange/ethedium bromide using fluorescence microscopy. Apoptosis percentage was measured by flow cytometric analysis using annexine-V-FITC. Also, DNA damage and bcl-2/bax were estimated by UV-method and western blot, respectively. RESULTS AND DISCUSSION: HPLC analysis revealed presence of three flavonoids, rutin, myricetin, and quercetin. MEZA showed satisfactory cytotoxicity in MTT assay (IC50 = 111.50 µg/ml). The extract significantly (p < 0.01) changed the tumor volume, viable, non-viable cell count, and hematological parameters towards the normal. Apoptotic activity of MEZA was confirmed by acridine orange/ethidium bromide staining, annexin-V-FITC staining, DNA fragmentation, and Bcl-2/Bax ratio. CONCLUSION: The study showed that MEZA has antitumor activity which may be due to the presence of flavonoids in the extract.

Antitumor Potential of Thevetia peruviana on Ehrlich's Ascites Carcinoma-Bearing Mice.

Methanol extracts of Thevetia peruviana (METP) (Apocynaceae) fruit showed antitumor activity against Ehrlich's ascites carcinoma (EAC) cell line in Swiss albino mice. The METP-treated group's tumor volume, tumor weight, and viable cell count were decreased compared to the EAC control group. Tumor volumes were 3.62±0.12 ml, 2.88±0.23 ml, and 1.34±0.17 ml for the EAC control group and the METP-treated groups (50 mg/kg and 100 mg/kg body weight), respectively. Nonviable cell counts were 4.44%±0.42%, 18.57%±3.07%, and 68.12%±5.32% in the EAC control group and the METP-treated groups (50 mg/kg and 100 mg/kg body weight), respectively. METP-treated EAC cell-bearing mice had an increased life span (48.69% and 83.78%) compared to the EAC control group. Hematological and serum biochemical profiles were restored to normal levels in METP-treated mice compared to the EAC control group. METP significantly (P<0.001) decreased lipid peroxidation and recovered reduced glutathione, superoxide dismutase, and catalase toward normal levels compared to the EAC control group. In summary, METP exhibited remarkable antitumor activity in Swiss albino mice, which is plausibly attributable to its augmentation of endogenous antioxidant mechanisms.

Oral delivery of therapeutic proteins and peptides: a review on recent developments.

Advent of recombinant technology in protein synthesis has given birth to a new range of biopharmaceuticals. These therapeutic peptides and proteins are now emerging as an imperative part of various treatment protocols especially in the cancer therapeutics. Despite extensive research efforts, oral delivery of therapeutic peptide or protein is still a challenge for pharmaceutical industries and researchers. Number of factors including high proteolytic activity and low pH conditions of gastrointestinal tract act as major barriers in the successful delivery of intact protein/peptide to the targeted site. Low permeability of protein/peptide across the intestinal barrier is also a factor adding to the low bioavailability. Therefore, because of the short circulatory half-life exhibited by peptides in vivo, they need to be administered frequently resulting in increased cost of treatment and low patient compliance. Nano-carrier-based delivery presents an appropriate choice of drug carriers owing to their property to protect proteins from degradation by the low pH conditions in stomach or by the proteolytic enzymes in the gastrointestinal tract. This review focuses on recent aspects and patents on oral delivery of therapeutic proteins and peptides with special emphasis on nano-carrier-based approach.