Robust single nucleus RNA sequencing reveals depot-specific cell population dynamics in adipose tissue remodeling during obesity.

Single nucleus RNA sequencing (snRNA-seq), an alternative to single cell RNA sequencing (scRNA-seq), encounters technical challenges in obtaining high-quality nuclei and RNA, persistently hindering its applications. Here, we present a robust technique for isolating nuclei across various tissue types, remarkably enhancing snRNA-seq data quality. Employing this approach, we comprehensively characterize the depot-dependent cellular dynamics of various cell types underlying adipose tissue remodeling during obesity. By integrating bulk nuclear RNA-seq from adipocyte nuclei of different sizes, we identify distinct adipocyte subpopulations categorized by size and functionality. These subpopulations follow two divergent trajectories, adaptive and pathological, with their prevalence varying by depot. Specifically, we identify a key molecular feature of dysfunctional hypertrophic adipocytes, a global shutdown in gene expression, along with elevated stress and inflammatory responses. Furthermore, our differential gene expression analysis reveals distinct contributions of adipocyte subpopulations to the overall pathophysiology of adipose tissue. Our study establishes a robust snRNA-seq method, providing novel insights into the mechanisms orchestrating adipose tissue remodeling during obesity, with broader applicability across diverse biological systems.

Liver-Derived S100A6 Propels ß-Cell Dysfunction in NAFLD.

Nonalcoholic fatty liver disease (NAFLD) is an independent predictor of systemic insulin resistance and type 2 diabetes mellitus (T2DM). However, converse correlates between excess liver fat content and ß-cell function remain equivocal. Specifically, how the accumulation of liver fat consequent to the enhanced de novo lipogenesis (DNL) leads to pancreatic ß-cell failure and eventually to T2DM is elusive. Here, we have identified that low-molecular-weight calcium-binding protein S100A6, or calcyclin, inhibits glucose-stimulated insulin secretion (GSIS) from ß cells through activation of the receptor for the advanced glycation end products and diminution of mitochondrial respiration. Serum S100A6 level is elevated both in human patients with NAFLD and in a high-fat diet-induced mouse model of NAFLD. Although serum S100A6 levels are negatively associated with ß-cell insulin secretory capacity in human patients, depletion of hepatic S100A6 improves GSIS and glycemia in mice, suggesting that S100A6 contributes to the pathophysiology of diabetes in NAFLD. Moreover, transcriptional induction of hepatic S100A6 is driven by the potent regulator of DNL, carbohydrate response element-binding protein (ChREBP), and ectopic expression of ChREBP in the liver suppresses GSIS in a S100A6-sensitive manner. Together, these data suggest elevated serum levels of S100A6 may serve as a biomarker in identifying patients with NAFLD with a heightened risk of developing ß-cell dysfunction. Overall, our data implicate S100A6 as, to our knowledge, a hitherto unknown hepatokine to be activated by ChREBP and that participates in the hepato-pancreatic communication to impair insulin secretion and drive the development of T2DM in NAFLD.

Distinct pathoclinical clusters among patients with uncontrolled type 2 diabetes: results from a prospective study in rural India.

INTRODUCTION: Inadequate glycemic control among patients with type 2 diabetes mellitus (T2DM) poses an enormous challenge. Whether this uncontrolled T2DM population is a heterogenous mix of disease subtypes remains unknown. Identification of these subtypes would result in a customized T2DM management protocol thereby paving the way toward personalized therapy. RESEARCH DESIGN AND METHODS: Electronic health records of 339 patients with uncontrolled T2DM patients followed up for a median period of 14 months were analyzed using Uniform Manifold Approximation and Projection followed by density-based spatial clustering of applications with noise. Baseline clinical features and final diagnoses with drug combinations were selected in the analysis. A 30 min oral glucose tolerance test was next performed for assessing the underlying insulin resistance and ß cell dysfunction. RESULTS: Three major clusters were identified. The first cluster characterized by recent onset T2DM had moderately preserved ß cell function. The second cluster with a longer duration of T2DM and associated hypertension showed the best glycemic control with dual antidiabetic therapy. The third cluster with the longest history of T2DM and no history of hypertension had the worst glycemic control in spite of the highest percentage of patients on triple therapy (34.58%) and quadruple therapy (8.41%). CONCLUSIONS: Uncontrolled T2DM comprises a heterogeneous population with respect to disease duration, presence of co-morbidities and ß cell function without significant difference in insulin resistance. Stratifying them on the basis of pathoclinical features is the first step toward a personalized management in T2DM.

Proteasome dysfunction under compromised redox metabolism dictates liver injury in NASH through ASK1/PPARγ binodal complementary modules.

Incidence of hepatotoxicity following acute drug-induced proteasomal inhibition and development of chronic proteasome dysfunction in obesity and insulin resistance underscores the crucial importance of hepatic protein homeostasis albeit with an elusive molecular basis and therapeutic opportunities. Apart from lipotoxicity and endoplasmic reticulum (ER) stress, herein we report that hepatocytes are highly susceptible to proteasome-associated metabolic stress attune to altered redox homeostasis. Bortezomib-induced proteasomal inhibition caused severe hepatocellular injury independent of ER stress via proapoptotic Apoptosis Signal-regulating Kinase 1 (ASK1)- c-Jun N-terminal kinase (JNK1)- p38 signaling concomitant with inadequate peroxisome proliferator-activated receptor γ (PPARγ)- Nuclear factor erythroid 2-related factor 2 (Nrf2) -driven antioxidant response. Although inhibition of ASK1 rescued acute hepatotoxicity, hepatic depletion of PPARγ or its physiological activator pigment epithelium-derived factor (PEDF) further aggravated liver injury even under ASK1 inhibition, emphasizing that endogenous PPARγ driven antioxidant activity serves as a prerequisite for the favorable therapeutic outcome of ASK1 inhibition. Consequently, ASK1 inhibitor selonsertib and PPARγ agonist pioglitazone in pharmacological synergism ameliorated bortezomib-induced hepatotoxicity and significantly prolonged survival duration in mice. Moreover, we showed that proteasome dysfunction is associated with ASK1 activation and insufficient PPARγ/Nrf2-driven antioxidative response in a subset of human nonalcoholic steatohepatitis (NASH) patients and the preclinical NASH model. The latter remains highly responsive to the drug combination marked by revamped proteasomal activity and alleviated hallmarks of NASH such as steatosis, fibrosis, and hepatocellular death. We thus uncovered a pharmacologically amenable interdependent binodal molecular circuit underlying hepatic proteasomal dysfunction and associated oxidative injury.

Selective aldehyde reductions in neutral water catalysed by encapsulation in a supramolecular cage.

The enhancement of reactivity inside supramolecular coordination cages has many analogies to the mode of action of enzymes, and continues to inspire the design of new catalysts for a range of reactions. However, despite being a near-ubiquitous class of reactions in organic chemistry, enhancement of the reduction of carbonyls to their corresponding alcohols remains very much underexplored in supramolecular coordination cages. Herein, we show that encapsulation of small aromatic aldehydes inside a supramolecular coordination cage allows the reduction of these aldehydes with the mild reducing agent sodium cyanoborohydride to proceed with high selectivity (ketones and esters are not reduced) and in good yields. In the absence of the cage, low pH conditions are essential for any appreciable conversion of the aldehydes to the alcohols. In contrast, the specific microenvironment inside the cage allows this reaction to proceed in bulk solution that is pH-neutral, or even basic. We propose that the cage acts to stabilise the protonated oxocarbenium ion reaction intermediates (enhancing aldehyde reactivity) whilst simultaneously favouring the encapsulation and reduction of smaller aldehydes (which fit more easily inside the cage). Such dual action (enhancement of reactivity and size-selectivity) is reminiscent of the mode of operation of natural enzymes and highlights the tremendous promise of cage architectures as selective catalysts.

Photochemical control of bacterial gene expression based on trans encoded genetic switches.

Controlling gene expression by light with fine spatiotemporal resolution not only allows understanding and manipulating fundamental biological processes but also fuels the development of novel therapeutic strategies. In complement to exploiting optogenetic tools, photochemical strategies mostly rely on the incorporation of photo-responsive small molecules into the corresponding biomacromolecular scaffolds. Therefore, generally large synthetic effort is required and the switching of gene expression in both directions within a single system remains a challenge. Here, we report a trans encoded ribo-switch, which consists of an engineered tRNA mimicking structure (TMS), under control of small photo-switchable signalling molecules. The signalling molecules consist of two amino glycoside molecules that are connected via an azobenzene unit. The light responsiveness of our system originates from the photo-switchable noncovalent interactions between the signalling molecule and the TMS switch, leading to the demonstration of photochemically controlled expression of two different genes. We believe that this modular design will provide a powerful platform for controlling the expression of other functional proteins with high spatiotemporal resolution employing light as a stimulus.

Synthesis, spectroscopic, electrochemical and photophysical properties of high band gap polymers for potential applications in semi-transparent solar cells.

BACKGROUND: The design of new polymers able to filter the electromagnetic spectrum and absorb distinctly in the UV and high-energy part of visible spectrum is crucial for the development of semi-transparent solar cells. Herein, we report on the synthesis and spectroscopic, electrochemical, and photophysical characteristics of three new polymers, namely (i) Poly(triamterene-co-terephthalate), (ii) Poly[triamterene-co- 3-(2-pyridyl)-5,6-diphenyl-1,2,4-triazine-p,p'-disulfonamide], and (iii) Poly(5-hydroxyindole-2-carboxylate) that might show promise as materials for semi-transparent solar cells. RESULTS: The energy band gap, refractive index, dielectric constant, and optical conductivity of the electron donor polymer, poly(triamterene-co-terephthalate), were determined to be 2.92 eV, 1.56, 2.44 and 2.43 × 104 S cm-1, respectively. The synthesized electron acceptor polymers showed a relatively high refractive index, dielectric constant, and optical conductivity. The presence of a direct allowed transition was confirmed between intermolecular energy bands of the polymers. CONCLUSIONS: The polymers showed relatively high energy gap and deep HOMO levels, making them strong absorbers of photons in the UV region and high energy part of the visible region. The synthesized donor and acceptors performed well relative to P3HT and fullerenes due to the close match of the HOMO and LUMO levels. With further development, the polymers could be viable for use as the active layers of semi-transparent solar cells.

Behavioral discrimination and time-series phenotyping of birdsong performance.

Variation in the acoustic structure of vocal signals is important to communicate social information. However, relatively little is known about the features that receivers extract to decipher relevant social information. Here, we took an expansive, bottom-up approach to delineate the feature space that could be important for processing social information in zebra finch song. Using operant techniques, we discovered that female zebra finches can consistently discriminate brief song phrases ("motifs") from different social contexts. We then applied machine learning algorithms to classify motifs based on thousands of time-series features and to uncover acoustic features for motif discrimination. In addition to highlighting classic acoustic features, the resulting algorithm revealed novel features for song discrimination, for example, measures of time irreversibility (i.e., the degree to which the statistical properties of the actual and time-reversed signal differ). Moreover, the algorithm accurately predicted female performance on individual motif exemplars. These data underscore and expand the promise of broad time-series phenotyping to acoustic analyses and social decision-making.

Deciphering the functional role of EGR1 in Prostaglandin F2 alpha induced luteal regression applying CRISPR in corpus luteum of buffalo.

BACKGROUND: PGF2α is essential for the induction of the corpus luteum regression which in turn reduces progesterone production. Early growth response (EGR) proteins are Cys2-His2-type zinc-finger transcription factor that are strongly linked to cellular proliferation, survival and apoptosis. Rapid elevation of EGR1 was observed after luteolytic dose of PGF2α. EGR1 is involved in the transactivation of many genes, including TGFß1, which plays an important role during luteal regression. METHODS: The current study was conducted in buffalo luteal cells with the aim to better understand the role of EGR1 in transactivation of TGFß1 during PGF2α induced luteal regression. Luteal cells from mid stage corpus luteum of buffalo were cultured and treated with different doses of PGF2α for different time durations. Relative expression of mRNAs encoding for enzymes within the progesterone biosynthetic pathway (3ßHSD, CYP11A1 and StAR); Caspase 3; AKT were analyzed to confirm the occurrence of luteolytic event. To determine if EGR1 is involved in the PGF2α induced luteal regression via induction of TGFß1 expression, we knocked out the EGR1 gene by using CRISPR/Cas9. RESULT: The present experiment determined whether EGR1 protein expression in luteal cells was responsive to PGF2α treatment. Quantification of EGR1 and TGFß1 mRNA showed significant up regulation in luteal cells of buffalo at 12 h post PGF2α induction. In order to validate the role of PGF2α on stimulating the expression of TGFß1 by an EGR1 dependent mechanism we knocked out EGR1. The EGR1 ablated luteal cells were stimulated with PGF2α and it was observed that EGR1 KO did not modulate the PGF2α induced expression of TGFß1. In PGF2α treated EGR1 KO luteal cell, the mRNA expression of Caspase 3 was significantly increased compared to PGF2α treated wild type luteal cells maintained for 12 h. We also studied the influence of EGR1 on steroidogenesis. The EGR1 KO luteal cells with PGF2α treatment showed no substantial difference either in the progesterone concentration or in StAR mRNA expression with PGF2α-treated wild type luteal cells. CONCLUSION: These results suggest that EGR1 signaling is not the only factor which plays a role in the regulation of PGF2α induced TGFß1 signaling for luteolysis.

Subcutaneous amyloidoma models for screening potential anti-fibrillating agents in vivo.

Here, we describe a robust protocol using mouse models to screen potential insulin-stabilizers and insulin moieties. We have generated a mouse model of amyloidoma, found in diabetic patients undergoing insulin therapy. This model can be used to screen potential insulin stabilizers and insulin moieties to prevent amyloidoma formation. This protocol can further be used for the preclinical validation of therapeutically relevant insulin stabilizers and formulations. The protocol highlights all the critical steps for generating amyloidoma in a preclinical model. For complete details on the use and execution of this profile, please refer to Mukherjee et al. (2021).

Deciphering the functional role of EGR1 in Prostaglandin F2 alpha induced luteal regression applying CRISPR in corpus luteum of buffalo

BACKGROUND: PGF2α is essential for the induction of the corpus luteum regression which in turn reduces progesterone production. Early growth response (EGR) proteins are Cys2-His2-type zinc-finger transcription factor that are strongly linked to cellular proliferation, survival and apoptosis. Rapid elevation of EGR1 was observed after luteolytic dose of PGF2α. EGR1 is involved in the transactivation of many genes, including TGFß1, which plays an important role during luteal regression. METHODS: The current study was conducted in buffalo luteal cells with the aim to better understand the role of EGR1 in transactivation of TGFß1 during PGF2α induced luteal regression. Luteal cells from mid stage corpus luteum of buffalo were cultured and treated with different doses of PGF2α for different time durations. Relative expression of mRNAs encoding for enzymes within the progesterone biosynthetic pathway (3ßHSD, CYP11A1 and StAR); Caspase 3; AKT were analyzed to confirm the occurrence of luteolytic event. To determine if EGR1 is involved in the PGF2α induced luteal regression via induction of TGFß1 expression, we knocked out the EGR1 gene by using CRISPR/Cas9. RESULT: The present experiment determined whether EGR1 protein expression in luteal cells was responsive to PGF2α treatment. Quantification of EGR1 and TGFß1 mRNA showed significant up regulation in luteal cells of buffalo at 12 h post PGF2α induction. In order to validate the role of PGF2α on stimulating the expression of TGFß1 by an EGR1 dependent mechanism we knocked out EGR1. The EGR1 ablated luteal cells were stimulated with PGF2α and it was observed that EGR1 KO did not modulate the PGF2α induced expression of TGFß1. In PGF2α treated EGR1 KO luteal cell, the mRNA expression of Caspase 3 was significantly increased compared to PGF2α treated wild type luteal cells maintained for 12 h. We also studied the influence of EGR1 on steroidogenesis. The EGR1 KO luteal cells with PGF2α treatment showed no substantial difference either in the progesterone concentration or in StAR mRNA expression with PGF2α-treated wild type luteal cells. CONCLUSION: These results suggest that EGR1 signaling is not the only factor which plays a role in the regulation of PGF2α induced TGFß1 signaling for luteolysis.

Modular and Versatile Trans-Encoded Genetic Switches.

Current bacterial RNA switches suffer from lack of versatile inputs and are difficult to engineer. We present versatile and modular RNA switches that are trans-encoded and based on tRNA-mimicking structures (TMSs). These switches provide a high degree of freedom for reengineering and can thus be designed to accept a wide range of inputs, including RNA, small molecules, and proteins. This powerful approach enables control of the translation of protein expression from plasmid and genome DNA.

Bis-Cyclometalated Iridium Complexes Containing 4,4'-Bis(phosphonomethyl)-2,2'-bipyridine Ligands: Photophysics, Electrochemistry, and High-Voltage Dye-Sensitized Solar Cells.

In this report, the synthesis and characterization of two bis-cyclometalated iridium(III) complexes are presented. Single-crystal X-ray diffraction shows that [Ir(ppy)2(4,4'-bis(diethylphosphonomethyl)-2,2'-bipyridine)]PF6 adopts a pseudooctahedral geometry. The complexes have an absorption feature in the near-visible-UV region and emit green light with excited-state lifetimes in hundreds of nanoseconds. The redox properties of these complexes show reversible behavior for both oxidative and reductive events. [Ir(ppy)2(4,4'-bis(phosphonomethyl)-2,2'-bipyridine)]PF6 readily binds to metal oxide supports, like nanostructured SnIV-doped In2O3 and TiO2, while still retaining reversible redox chemistry. When incorporated as the photoanode in dye-sensitized solar cells, the devices exhibit open-circuit voltages of >1 V, which is a testament to their strength of these iridium(III) complexes as photochemical oxidants.

Tunable Redox Potential, Optical Properties, and Enhanced Stability of Modified Ferrocene-Based Complexes.

We report a series of ferrocene-based derivatives and their corresponding oxidized forms in which the introduction of simple electron donating groups like methyl or tert-butyl units on cyclopentadienyl-rings afford great tunability of Fe+III/Fe+II redox potentials from +0.403 V down to -0.096 V versus saturated calomel electrode. The spin forbidden d-d transitions of ferrocene derivatives shift slightly toward the blue region with an increasing number of electron-donating groups on the cyclopentadienyl-rings with very little change in absorptivity values, whereas the ligand-to-metal transitions of the corresponding ferricinium salts move significantly to the near-IR region. The electron-donating groups also contribute in the strengthening of electron density of Fe+III d-orbitals, which therefore improves the chemical stability against the oxygen reaction. Further, density functional theory calculations show a reducing trend in outer shell reorganization energy with an increasing number of the electron donating units.

Transcriptional Regulation of Thrombospondins and Its Functional Validation through CRISPR/Cas9 Mediated Gene Editing in Corpus Luteum of Water Buffalo (Bubalus Bubalis).

BACKGROUND/AIMS: Thrombospondins (TSPs) are large multi-modular proteins, identified as natural angiogenesis inhibitors that exert their activity by binding to CD36 and CD47 receptors. The anti-angiogenic effect of TSPs in luteal regression of water buffalo has not been addressed. The present study characterized the expression pattern and localization of TSPs and their receptors in ovarian corpus luteum during different stages of development in buffalo. This study also elucidated the effect of exogenous Thrombospondin1 (TSP1) or the knocking out of the endogenous protein on luteal cell viability and function. Further, the in vitro transcriptional interaction of TSP1 with hormones, LH, PGF2α and angiogenic growth factors, VEGF and FGF2 were also evaluated. METHODS: First, the CLs were classified into four groups based on macroscopic observation and progesterone concentration. mRNA expression of examined factors was measured by qPCR, localization by immunoblotting and immunohistochemistry. TSP1 was knocked out (KO) in cultured luteal cells isolated from late luteal stage CLs (day 1116) by CRISPR/Cas9 mediated gene editing technology in order to functionally validate the TSP1 gene. Isolated cells from late stage CLs were also stimulated with different doses of TSP1, LH, PGF2α, VEGF and FGF2 for various time intervals to determine transcriptional regulation of thrombospondins. RESULTS: mRNA expression of TSPs and their receptors were found to be significantly higher in late and regressed stage of CL as compared to other groups which was consistent with the findings of immunoblotting and immunolocalization experiments. It was observed that TSP1 induced apoptosis, down regulated angiogenic growth factors, VEGF and FGF2 and attenuated progesterone production in cultured luteal cells. However, knocking out of endogenous TSP1 with CRISPR/Cas9 system improved the viability of luteal cells, progesterone synthesis and upregulated the expression of VEGF and FGF2 in the KO luteal cells. PGF2α induced the upregulation of TSPs and Caspase 3 transcripts, whereas treatment with LH and angiogenic growth factors (VEGF and FGF2) down regulated the TSP system in luteal cells. CONCLUSION: Collectively, these data provide evidence that thrombospondins along with their receptors are expressed at varying levels in different stages of CL progression with maximum expression during the late and regressing stages. These results are consistent with the hypothesis that thrombospondins stimulated by PGF2α plays an essential modulatory role in bringing about structural and functional luteolysis in buffalo.

Regulation of steroidogenic function of luteal cells by thrombospondin and insulin in water buffalo (Bubalus bubalis).

The present study examined the effect of exogenous thrombospondin 1 (TSP1) on the steroidogenic function of luteal cells cultured invitro. Furthermore, the transcriptional interaction of insulin with TSP1 and its receptor, cluster of differentiation 36 (CD36) were also investigated. At the highest dose (500ngmL-1) TSP1 significantly downregulated the expression of the angiogenic marker von Willebrand factor (vWF) and progesterone production in cultured luteal cells. Moreover, the simultaneous upregulation in the expression of caspase 3 by exogenous TSP1 was consistent with a reduction in the number of viable luteal cells as determined by 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltertrazolium bromide (MTT) assay after 72h of culture. However, the expression of critical enzymes in the progesterone synthetic pathway was not significantly modulated by treatment with TSP1 in cultured luteal cells. Knocking out of endogenous TSP1 with the clustered regularly interspaced short palindromic repeats (CRISPR)/ CRISPRassociated protein9 (Cas9) system improved the viability of luteal cells as well as increasing progesterone production and decreasing caspase 3 activation. Insulin treatment suppressed the expression of TSP1 and CD36 in cultured luteal cells in a dose- and time-dependent manner. To conclude, TSP1 acts as a negative endogenous regulator of angiogenesis that attenuates progesterone production, possibly by reducing the number of luteal cells via apoptosis during luteal regression, whereas insulin as a luteinising signal may have inhibited the thrombospondin system for the efficient development of luteal function.

Luteolin attenuates acute lung injury in experimental mouse model of sepsis.

The present investigation was undertaken to assess the result of pretreatment of luteolin in sepsis-induced acute lung injury in mice and its mechanism of action. Luteolin was administered intraperitoneally one hour before caecal ligation and puncture (CLP) surgery in mice. Acute lung injury was assessed by estimation of different parameters like lung edema, protein content, cytokines level, oxidative stress, inducible nitric oxide synthase (iNOS), intercellular adhesion molecule (ICAM)-1 expression and histopathology. Pretreatment of mice with luteolin showed decrease lung edema and protein content in tissue and bronchoalveolar lavage fluid (BALF). However, mice pretreated with luteolin showed reduction (p = 0.92) in blood and lung tissue bacterial counts however it was non significant. Further, luteolin showed significant reduction in interleukin (IL)-6 and IL-1ß in lung tissue which are the proinflammatory cytokines. However, plasma IL-1ß and tissue tumor necrosis factor (TNF)-α level decrease (p = 0.24; p = 0.19) with this pretreatment. Further, ICAM-1 mRNA expression and nuclear factor (NF)-kappa B protein expression were significantly (p < 0.01) decreased in luteolin pretreated septic mice. The lung iNOS level, iNOS mRNA and protein expressions were markedly (p = 0.25; p = 0.50; p = 0.06) altered with luteolin pretreatment, respectively. Also, significant reduction in lipid peroxidation and increase in the activity of antioxidant enzymes like superoxide dismutase (SOD) and catalase was noted with luteolin pretreatment. However, luteolin did not alter (p = 0.36) the non enzymatic antioxidant GSH activity in septic mice. Histopathology of lung tissue showed reduction in lung injury with the luteolin pretreatment in septic mice. The study suggests that luteolin showed attenuation in sepsis-induced acute lung injury in mice through suppression in ICAM-1, NF-kappa B, oxidative stress and partially iNOS pathways.

Kaempferol attenuates acute lung injury in caecal ligation and puncture model of sepsis in mice.

AIM OF THE STUDY: Kaempferol is a flavonoid and important part of the diet. Kaempferol has shown antioxidant, antiinflammatory and antidiabetic activities in various studies. However, protective potential of kaempferol in acute lung injury induced by sepsis and its mechanism remains unclear. The present study was undertaken to evaluate the effect of kaempferol in sepsis-induced acute lung injury in mice and its possible mechanism of action. MATERIALS AND METHODS: Acute lung injury was induced by CLP surgery in mice. Kaempferol (100 mg/kg bw) was administered orally one hour before caecal ligation and puncture surgery in mice. Mice were divided into four groups sham, KEM+sham, sepsis (CLP), and KEM+sepsis. Assessment of lung injury was done by estimation of protein content in lung tissue, lung edema, proinflammatory cytokines in plasma and lung tissue, oxidative stress, antioxidant enzymes, nitrite production, and histopathology. RESULTS: Kaempferol pretreated mice showed significant (P < 0.001) decrease in water content in lungs. Kaempferol pretreatment showed reduction in cytokines IL-6, IL-1ß, and TNF-α in plasma as well as in lung tissue in comparison with septic mice without pretreatment. Pretreatment with kaempferol did not show any reduction in MDA level in comparison with septic mice. Antioxidant enzymes SOD and catalase and nonenzymatic antioxidant GSH activities were also increased with kaempferol pretreatment in septic mice. Further, kaempferol pretreatment reduced the lung tissue nitrite level (P < 0.01) and iNOS (P < 0.05) level in septic mice. A significant (P < 0.01) downregulation of mRNA expression of ICAM-1 and iNOS was observed with this pretreatment. Kaempferol pretreatment did not decrease bacterial load in septic mice. Mice pretreated with kaempferol followed by sepsis showed lesser infiltration of cells and more arranged alveolar structure in histopathological analysis. CONCLUSIONS: The study suggests that kaempferol showed attenuation in sepsis-induced acute lung injury in mice through suppression of oxidative stress, iNOS, and ICAM-1 pathways.

Impact of oocyte-secreted factors on its developmental competence in buffalo.

Oocyte-secreted factors (OSFs) play an important role in the acquisition of oocyte developmental competence through bidirectional cross-talk between oocyte and cumulus cells via gap junctions. Thus, the present study was designed to investigate the effect of two OSFs, growth differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15), on the developmental competence of buffalo oocytes derived from two different follicle sizes. Cumulus-oocyte complexes (COCs) from large follicles (LF, >6 mm) or small follicles (SF, 0.05) between DOs and combination groups. Relative mRNA analysis revealed significantly higher (P > 0.05) expression of the cumulus cell marker genes EGFR, HAS2, and CD44 in LF-derived than SF-derived oocyte; the expression of these markers was significantly higher (P > 0.05) in DOs and combination groups, irrespective of the follicle size. These results suggested that LF-derived oocytes have a higher developmental competence than SF-derived oocytes and that supplementation of GDF9 and BMP15 modulates the developmental competence of buffalo oocytes by increasing the relative abundance of cumulus-enabling factors and thereby increasing cleavage and the quality of blastocyst production.

Orientation and Incorporation of Photosystem I in Bioelectronics Devices Enabled by Phage Display.

Interfacing proteins with electrode surfaces is important for the field of bioelectronics. Here, a general concept based on phage display is presented to evolve small peptide binders for immobilizing and orienting large protein complexes on semiconducting substrates. Employing this method, photosystem I is incorporated into solid-state biophotovoltaic cells.