Identification, crystallization, and first X-ray structure analyses of phenyl boronic acid-based inhibitors of human carbonic anhydrase-II.

Human carbonic anhydrases (hCAs) play a central role in various physiological processes in the human body. HCAs catalyze the reversible hydration of CO2 into HCO3-, and hence maintains the fluid and pH balance. Overexpression of CA II is associated with diseases, such as glaucoma, and epilepsy. Therefore, CAs are important clinical targets and inhibition of different isoforms, especially hCA II is used in treatment of glaucoma, altitude sickness, and epilepsy. Therapeutically used CA inhibitors (CAI) are sulfonamide-based, such as acetazolamide, dichlorphenamide, methazolamide, ethoxzolamide, etc. However, they exhibit several undesirable effects such as numbness, tingling of extremities, malaise, metallic taste, fatigue, renal calculi, and metabolic acidosis. Therefore, there is an urgent need to identify safe and effective inhibitors of the hCAs. In this study, different phenyl boronic acids 1-5 were evaluated against bovine (bCA II) and hCA II. Among all, compound 1 (4-acetylphenyl boronic acid) was found to be active against bCAII and hCA II with IC50 values of 246 ± 0.48 and 281.40 ± 2.8 µM, respectively, while the remaining compounds were found in-active. Compound 1 was identified as competitive inhibitor of hCA II enzyme (Ki = 283.7 ± 0.002 µM). Additionally, compound 1 was found to be non-toxic against BJ Human fibroblast cell line. The X-ray crystal structure for hCA II in-complex with compound 1 was evaluated to a resolution of 2.6 Å. In fact, this the first structural analysis of a phenyl boron-based inhibitor bound to hCA II, allowing an additional structure-activity analysis of the compounds. Compound 1 was found to be directly bound in the active site of hCA II by interacting with His94, His119, and Thr199 residues. In addition, a bond of 3.11 Å between the zinc ion and coordinated boron atom of the boronic acid moiety of compound 1 was also observed, contributing to binding affinity of compound 1 for hCA II. PDB ID: 8IGF.

Comparison of Honey & Povidone Iodine dressings in Post-Cesarean Surgical Site Wound Infection Healing.

Objective: To compare the healing of post-cesarean infected wounds by a combination of honey and povidone iodine & povidone-iodine alone dressings using ASEPSIS Score. Method: This prospective randomised cohort study was carried out at at OBGYN Department Jinnah Postgraduate Medical Center from 1st August 2022 till 31st December 2022. Patients were randomly allocated into groups A and B. Group-A was dressed with honey povidone-iodine, while Group-B only had povidone-iodine. The ASEPSIS score was calculated in both groups on day fifth, 7th, 9th, and 10th day. Patients were followed till full recovery of the wound. Results: A total of 70 women were included and equally allocated into two groups A & B (35 each). On day five mean ASEPSIS score in Group-A was 36.14 and that in Group- B was 37.74. No significant difference in scores were noted on day five [t (68) = -.753, p = .454] & day seven Group-A 28.63 vs Group-B 32.11 [t (68) = -1.302, p = .197]. Significant improvements in ASEPSIS scores were noted on days ninth & tenth. On day nine ASEPSIS score of Group-A was 21.54 and that of Group-B was 27.14 [t (68) = -2.056, p = .044]. On day tenth the mean ASEPSIS score of Group-A was 18.26, while that of Group-B was 23.86 [t (68) = -2.021, p = .047]. The mean time required for the wound to heal in Group-A and Group-B was eighteen & twenty-one days respectively. Conclusion: Significant improvements in ASEPSIS score occurred with the use of honey with povidone-iodine combination.

Simple two-step purification and characterisation of peroxidase from Citrullus colocynthis.

Peroxidase is a biotechnologically important enzyme. The purification of peroxidase from the root of Citrullus colocynthis was carried out in a simple two-step process with maximum purity level. The sample was extracted in a high salt buffer, and the enzyme was partially purified with a Q-Sepharose anion exchange column. Final purification was carried out with HighLoad 16/600 Superdex G-75 column. The purified protein was analysed with SDS gel electrophoresis, which suggested a single band of approximately 35 kDa. Further, the enzyme was identified with the help of Mass spectrometric analysis using an ESI-QTOF Mass spectrometer. The study will be helpful for the isolation and its commercial uses in biotechnology.

Comparative evaluation of ethyl acetate and n-Hexane extracts of Cannabis sativa L. leaves for muscle function restoration after peripheral nerve lesion.

Peripheral nerve injuries are one of those complex medical conditions for which a highly effective first-line treatment is currently missing. The use of natural compound as medicines to treat various disorders has a long history. Our previous research explored that crude Cannabis sativa L. accelerated the recovery of sensorimotor functions following nerve injury. The purpose of the current study was to investigate the effects of n-Hexane and ethyl acetate extracts of C. sativa L. leaves on the muscle function restoration in a mouse model after sciatic nerve injury. For this purpose, albino mice (n = 18) were equally divided into control and two treatment groups. The control group was fed on a plain diet while treatment groups were given a diet having n-Hexane (treatment 1) and ethyl acetate (treatment 2) extracts of C. sativa L. (10 mg/kg body weight), respectively. The hot plate test (M = 15.61, SD = 2.61, p = .001), grip strength (M = 68.32, SD = 3.22, p < .001), and sciatic functional index (SFI) (M = 11.59, SD = 6.54, p = .012) assessment indicated significant amelioration in treatment 1 as compared to treatment 2 group. Furthermore, muscle fiber cross-sectional area revealed a noticeable improvement (M = 182,319, SD = 35.80, p = .013) in treatment 1 while muscle mass ratio of Gastrocnemius (M = 0.64, SD = 0.08, p = .427) and Tibialis anterior (M = 0.57, SD = 0.04, p = .209) indicated nonsignificant change. A prominent increase in total antioxidant capacity (TAC) (M = 3.76, SD = 0.38, p < .001) and momentous decrease in total oxidant status (TOS) (M = 11.28, SD = 5.71, p < .001) along with blood glucose level indicated significant difference (M = 105.5, SD = 9.12, p < 0.001) in treatment 1 group. These results suggest that treatment 1 has the ability to speed up functional recovery after a peripheral nerve lesion. Further research is necessary, nevertheless, to better understand the extract's actual curative properties and the mechanisms that improve functional restoration.

Insight into the structural basis of the dual inhibitory mode of Lima bean (Phaseolus lunatus) serine protease inhibitor.

Bovine pancreatic trypsin was crystallized, in-complex with Lima bean trypsin inhibitor (LBTI) (Phaseolus lunatus L.), in the form of a ternary complex. LBTI is a Bowman-Birk-type bifunctional serine protease inhibitor, which has two independent inhibitory loops. Both of the loops can inhibit trypsin, however, only the hydrophobic loop is specific for inhibiting chymotrypsin. The structure of trypsin incomplex with the LBTI has been solved and refined at 2.25 Å resolution, in the space group P41, with Rwork /Rfree values of 18.1/23.3. The two binding sites of LBTI differ in only two amino acids. Lysine and leucine are the key residues of the two different binding loops positioned at the P1, and involved in binding the S1 binding site of trypsin. The asymmetric unit cell contains two molecules of trypsin and one molecule of LBTI. The key interactions include hydrogen bonds between LBTI and active site residues of trypsin. The 3D structure of the enzyme-inhibitor complex provided details insight into the trypsin inhibition by LBTI. To the best of our knowledge, this is the first report on the structure of trypsin incomplex with LBTI.

Axonal Regeneration: Underlying Molecular Mechanisms and Potential Therapeutic Targets.

Axons in the peripheral nervous system have the ability to repair themselves after damage, whereas axons in the central nervous system are unable to do so. A common and important characteristic of damage to the spinal cord, brain, and peripheral nerves is the disruption of axonal regrowth. Interestingly, intrinsic growth factors play a significant role in the axonal regeneration of injured nerves. Various factors such as proteomic profile, microtubule stability, ribosomal location, and signalling pathways mark a line between the central and peripheral axons' capacity for self-renewal. Unfortunately, glial scar development, myelin-associated inhibitor molecules, lack of neurotrophic factors, and inflammatory reactions are among the factors that restrict axonal regeneration. Molecular pathways such as cAMP, MAPK, JAK/STAT, ATF3/CREB, BMP/SMAD, AKT/mTORC1/p70S6K, PI3K/AKT, GSK-3ß/CLASP, BDNF/Trk, Ras/ERK, integrin/FAK, RhoA/ROCK/LIMK, and POSTN/integrin are activated after nerve injury and are considered significant players in axonal regeneration. In addition to the aforementioned pathways, growth factors, microRNAs, and astrocytes are also commendable participants in regeneration. In this review, we discuss the detailed mechanism of each pathway along with key players that can be potentially valuable targets to help achieve quick axonal healing. We also identify the prospective targets that could help close knowledge gaps in the molecular pathways underlying regeneration and shed light on the creation of more powerful strategies to encourage axonal regeneration after nervous system injury.

Vitamin C supplementation ameliorates liver function profile and antiviral treatment response in Hepatitis C patients.

An imbalance between oxidative stress and antioxidative defence mediates a variety of diseases pathogenesis. The present study aims to assess the possible outcome of supplementation of oral vitamin-C (VC), an antioxidant, in Viral Hepatitis C (HCV) treatment as an adjuvant therapy. 200 HCV-patients were selected, 100 were given Vitamin-C (1000 mg/day) along with anti HCV treatment (sofosbuvir plus daclatasvir) while the other 100 took only anti-HCV treatment for 4weeks. The serum ascorbic acid (Vitamin-C) levels and functions of the liver were tested before and after the VC supplementation. HCV patients with relatively low serum ascorbic acid showed significant improvement after the intake of vitamin C. After 4 weeks of treatment, AST, ALP, albumin, and total, direct and indirect bilirubin were improved significantly in the VC group; whereas only ALT and indirect bilirubin were improved in both groups when associated with the control subjects. Comparing the two treatment groups at 4weeks; more effective and significant improvement was observed in ALT (p<0.01), AST (p<0.001), direct (p<0.01) and indirect bilirubin (p<0.001), total proteins (p<0.001) and albumin (p<0.05) in patients with VC supplementation on anti-viral treatment compared to only anti-viral treatment group. Thus, VC supplementation improves the antiviral therapy outcome by bestowing a beneficial effect in minimizing liver damage in HCV cases.

Synthesis, anticancer evaluation and molecular docking studies of methotrexate's novel Schiff base derivatives against malignant glioma cell lines.

Recent years have witnessed advancement in cancer research that has led to the development of improved cytotoxic therapies with reduced side effects. Methotrexate (MTX) is a commonly used anticancer drug having robust activity, but with serious side effects. Several derivatives of MTX have been reported by modification at different sites to reduce its side effects and enhance efficacy. The current work describes the development of active MTX Schiff base derivatives by treating MTX with several aldehydes viz 2-chlorobenzaldehyde, 3-nitrobenzaldehyde, 5-chloro-2-hydroxybenz-aldehyde, 2-hydroxy-5-nitrobenzaldehyde, 2-thiocarboxyaldehyde, trans-2-pentenal and glutaraldehyde. Newly synthesized derivatives were evaluated for their anticancer potential against human malignant glioma U87 (MG-U87) cell lines at different concentrations of 200 µM, 100 µM, 50 µM, 25 µM, 12.5 µm, 6.25 µm and 0 µM. MTX derivatives with 2-Chlorobenzaldehyde (IC50 ∼100 µM), 2-Thiocarboxyaldehyde (IC50 <200 µM) and 2- Pentenal (IC50 ∼250 µM) showed much better activity at 100 µM compared to 400 µM concentration of MTX. Molecular docking studies were performed that showed a good correlation with the results obtained from in vitro experiments. The excellent agreement between molecular modeling and growth inhibition assay shows that the binding mode hypothesis is justly close to the experimentally biological values, therefore, may prove helpful for further lead optimization and clinical trials.Communicated by Ramaswamy H. Sarma.

Natural products based crown ethers: synthesis and their anticancer potential.

Natural products based novel crown ethers have been prepared by employing biologically active natural structures including tetrahydroisoquinoline, chrysin and biochanin-A as the side arms. The resulting crown scaffolds were evaluated for their anticancer potential against two cancer cell lines i.e. NCI-H460 (non-small lung carcinoma), MCF-7 (breast adenocarcinoma). The comparative study showed that the addition of crown scaffold put marked effects on antiproliferative profile of parent natural precursors and is significant for lung carcinoma in particular. Biochanin-A derived crown ether showed three (03) folds higher antiproliferative activity (IC50 = 6.08 ± 0.07 µM) against lung carcinoma as compared to standard drug cisplatin (IC50 = 19.00 ± 1.24 µM). Cytotoxic trends for NIH-3T3 cell lines were also examined and found reduced as compared to parent natural structures. Hence, these findings could open a new pathway towards developing effective carcinostatic drugs.HIGHLIGHTSFour natural products based novel crown ethers have been developed.Comparative antiproliferative screening of crown ethers and natural precursors.Addition of crown showed marked effects on anticancer profile of natural products.Crown formation is significant for lung carcinoma potential in particular.Biochanin-A derived crown ether found three folds more active than standard drug.

Dietary biomolecules as promising regenerative agents for peripheral nerve injury: An emerging nutraceutical-based therapeutic approach.

Peripheral nerve damage is a debilitating condition that can result in partial or complete functional loss as a result of axonal degeneration, as well as lifelong dependence. Many therapies have been imbued with a plethora of positive features while posing little risks. It is worth noting that these biomolecules work by activating several intrinsic pathways that are known to be important in peripheral nerve regeneration. Although the underlying mechanism is used for accurate and speedy functional recovery, none of them are without side effects. As a result, it is believed that effective therapy is currently lacking. The dietary biomolecules-based intervention, among other ways, is appealing, safe, and effective. Upregulation of transcription factors, neurotrophic factors, and growth factors such as NGF, GDNF, BDNF, and CTNF may occur as a result of these substances' dietary intake. Upregulation of the signaling pathways ERK, JNK, p38, and PKA has also been seen, which aids in axonal regeneration. Although several mechanistic approaches to understanding their involvement have been suggested, more work is needed to reveal the amazing properties of these biomolecules. We have discussed in this article that how different dietary biomolecules can help with functional recovery and regeneration after an injury. PRACTICAL APPLICATIONS: Based on the information known to date, we may conclude that treatment techniques for peripheral nerve injury have downsides, such as complications, donor shortages, adverse effects, unaffordability, and a lack of precision in efficacy. These difficulties cast doubt on their efficacy and raise severe concerns about the prescription. In this situation, the need for safe and effective therapeutic techniques is unavoidable, and dietary biomolecules appear to be a safe, cost-efficient, and effective way to promote nerve regeneration following an injury. The information on these biomolecules has been summarized here. Upregulation of transcription factors, neurotrophic factors, and growth factors, such as NGF, GDNF, BDNF, and CTNF, as well as the ERK, JNK, p38, and PKA, signaling pathways, may stimulate axonal regeneration.

Dysregulated Autophagy Leads to Oxidative Stress and Aberrant Expression of ABC Transporters in Women with Early Miscarriage.

Early miscarriage (EMC) is a devastating obstetrical complication. ATP-binding cassette (ABC) transporters mediate cholesterol transfer across the placenta and enhance cell survival by effluxing substrates from target cells in the presence of stressors. Recent evidence reports an intricate interplay between autophagy and ABC transporters. We hypothesized that dysregulated autophagy and oxidative stress (OS) in the placenta leads to abnormal expression of membrane transporters contributing to poor pregnancy survival in EMC. We determined mRNA and protein expression of autophagy genes (Beclin-1/Bcl-2/LC3I/LC3II/p62) and ABC transporters (ABCA1/ABCG1/ABCG2) in placentae from EMC patients (n = 20), term controls (n = 19), first trimester (n = 6), and term controls (n = 5) controls. Oxidative/antioxidant status and biomarkers of oxidative damage were evaluated in maternal serum and placentae from EMC and healthy controls. In EMC, placental expression of LC3II/LC3I as well as of the key autophagy regulatory proteins Beclin-1 and Bcl-2 were reduced, whereas p62 was increased. Both in the serum and placentae of EMC patients, total OS was elevated reflected by increased oxidative damage markers (8-OHdG/malondialdehyde/carbonyl formation) accompanied by diminished levels of total antioxidant status, catalase, and total glutathione. Furthermore, we found reduced ABCG1 and increased ABCG2 expression. These findings suggest that a decreased autophagy status triggers Bcl-2-dependent OS leading to macromolecule damage in EMC placentae. The decreased expression of ABCG1 contributes to reduced cholesterol export to the growing fetus. Increasing ABCG2 expression could represent a protective feedback mechanism under inhibited autophagy conditions. In conclusion, dysregulated autophagy combined with increased oxidative toxicity and aberrant expression of placental ABC transporters affects materno-fetal health in EMC.

Sex-Specific Regulation of Interferon-γ Cytotoxicity in Mouse Liver by Autophagy.

BACKGROUND AND AIMS: Interferon-γ (IFNγ) is a central activator of immune responses in the liver and other organs. IFNγ triggers tissue injury and inflammation in immune diseases, which occur predominantly in females for unknown reasons. Recent findings that autophagy regulates hepatotoxicity from proinflammatory cytokines led to an examination of whether defective hepatocyte autophagy underlies sex-specific liver injury and inflammation induced by IFNγ. APPROACH AND RESULTS: A lentiviral autophagy-related 5 (Atg5) knockdown was performed to decrease autophagy-sensitized alpha mouse liver (AML 12) hepatocytes to death from IFNγ in combination with IL-1ß or TNF. Death was necrosis attributable to impaired energy homeostasis and adenosine triphosphate depletion. Male mice with decreased autophagy from a tamoxifen-inducible, hepatocyte-specific Atg5 knockout were resistant to IFNγ hepatotoxicity whereas female knockout mice developed liver injury and inflammation. Female mice had increased IFNγ-induced signal transducer and activator of transcription 1 (STAT1) levels compared to males. Blocking STAT1, but not interferon regulatory factor 1, signaling prevented IFNγ-induced hepatocyte death in autophagy-deficient AML12 cells and female mice. The mechanism of death is STAT1-induced overexpression of nitric oxide synthase 2 (NOS2) as in vitro hepatocyte death and in vivo liver injury were blocked by NOS2 inhibition. CONCLUSIONS: Decreased hepatocyte autophagy sensitizes mice to IFNγ-induced liver injury and inflammation through overactivation of STAT1 signaling that causes NOS2 overexpression. Hepatotoxicity is restricted to female mice, suggesting that sex-specific effects of defective autophagy may underlie the increased susceptibility of females to IFNγ-mediated immune diseases.

Isoflavanquinones from Abrus precatorius roots with their antiproliferative and anti-inflammatory effects.

Phytochemical studies on the root of Abrus precatorius Linn. (Fabaceae), leads towards the identification of four undescribed (abruquinones M, N, O, and P), and seven known abruquinones, (abruquinones A, E, B, F, I, D, and G). Spectroscopic analyses (1D, and 2D NMR, HRESI-MS) were used in elucidating structures of the all compounds. Evaluation of anticancer activities of the isolated isoflavanquinones revealed that abruquinones M, and N showed cytotoxicity against oral CAL-27 (IC50 values 6.48 and 5.26 µM, respectively), and colon (Caco-2) cell lines (IC50 values 15.79 and 10.33 µM, respectively). Abruquinone M also inhibited the growth of lung cancer cells (NCI-H460) with IC50 of 31.33 µM. The isolated isoflavanquiones also showed potent anti-inflammatory potential through phagocyte oxidative burst and pro-inflammatory cytokine TNF-α inhibition in vitro. These findings suggest isoflavanquinones from A. precatorius roots as candidates for further research in cancer treatment.

Methanolic extract of Fennel (Foeniculum vulgare) escalates functional restoration following a compression injury to the sciatic nerve in a mouse model.

Peripheral nerve injury (PNI) is one of the major health concerns faced by the community at present. Till now, available therapeutic approaches are ineffective to fully heal a nerve injury and to assure the functional recovery entirely. Natural compounds can prove attractive and effective drug candidates to bridge up this gap. In this scenario, the present study was designed to explore the role of methanolic extract of Foeniculum vulgare (F. vulgare) seeds in accelerating the function regain following a sciatic nerve injury in a mouse model. For this purpose, 12 adult healthy albino mice (BALB/C), 8-10 weeks old, were grouped as control (Ctrl, n = 6) and treatment (Trt, n = 6). The treated group was given methanolic extract of F. vulgare (200 mg/kg per day) started from the day of nerve crush until the end of the study. The sensorimotor function regain assessed by hot plate test, grip strength, and SFI assessments was found significantly (p < .05) ameliorated in the F. vulgare-treated group. A prominent improvement in the muscle mass of the treated group further affirmed these effects. Furthermore, morphometric analysis of muscle fiber cross-sectional area of tibialis anterior muscle between groups revealed a noticeable improvement in muscle fibers' diameter of the treated group. Conclusively, these findings suggest that F. vulgare methanolic extract exhibits the potential to escalate functional recovery following a peripheral nerve injury. However, the real players of this extract and the mechanism involved in boosting functional restoration need to be dissected by further work.

Moringa oleifera Lam.ameliorates the muscles function recovery following an induced insult to the Sciatic nerve in a mouse model.

Peripheral nerve injury (PNI) is an incapacitating situation and has no effective therapy until now. We examined the possible role of crude leaves of Moringa oleifera Lam. at 200 mg/kg body weight in accelerating the functional regain in the sciatic nerve lesion induced mouse model (Adult male albino mice (BALB/c). Motor functions were evaluated by using the sciatic functional index, muscle mass, and muscle grip strength measurement, whereas the sensory functions were evaluated by using the hot plate test. Blood glucose levels and blood cell composition were also analyzed. We found that the Moringa oleifera crude leaves endorse the sensory and motor functions reclamation following the PNI with a statistically significant difference (p < .05). It also revitalizes the gastrocnemius muscle by mass restoration with glycemic management perspective. Conclusively, the crude powder of Moringa oleifera leaves exhibited a function restoration boosting property and further detailed studies for its application as a therapeutic agent are strongly recommended.

Strychnos nux-vomica L. seed preparation promotes functional recovery and attenuates oxidative stress in a mouse model of sciatic nerve crush injury.

BACKGROUND: Peripheral nerve injury is a debilitating condition that may lead to partial or complete motor, sensory and autonomic function loss and lacks effective therapy until date. Therefore, it is quite imperative to explore impending remedies for rapid and accurate functional retrieval following such conditions. Natural product-based intervention can prove effective to facilitate the process of functions regain. METHODS: Here, we investigated the effect of processed Strychnos nux-vomica seeds at a dose of 250 mg/kg body weight in a mouse model of induced Sciatic nerve lesion in promoting the recovery of the functions. A compression injury was induced in the Sciatic nerve of the right leg in the mice. Sensory function recovery was evaluated by hot-plate and formalin tests, whereas the motor function retrieval was assessed by measuring muscle grip strength, sciatic functional index, and muscle mass restoration. Oxidative stress and blood cell count were measured by biochemistry and haematological analyses. RESULTS: This study indicates that Strychnos nux-vomica seeds enhance the rate of recovery of both sensory and motor functions. It helps restore the muscle mass, attenuates total oxidant status and enhances the total anti-oxidant capacity of the biological system. Moreover, the treated animals manifested an enhanced glucose tolerance aptitude and augmented granulocyte and platelet counts. Improved oxidant control, enhanced glucose sensitivity and amended granulocyte and platelet counts are likely to contribute to the advantageous effects of Strychnos nux-vomica, and warrant further in-depth studies for deciphering possible mechanisms and identification of active constituent(s) responsible for these effects. CONCLUSION: Strychnos nux-vomica seed offers functional recovery promoting effects following a mechanical injury to the Sciatic nerve and the possible reasons behind this effect can be reduced oxidative stress and improved glycaemic control. Further and detailed investigations can unravel this mystery.

Chemical activation of SAT1 corrects diet-induced metabolic syndrome.

The pharmacological targeting of polyamine metabolism is currently under the spotlight for its potential in the prevention and treatment of several age-associated disorders. Here, we report the finding that triethylenetetramine dihydrochloride (TETA), a copper-chelator agent that can be safely administered to patients for the long-term treatment of Wilson disease, exerts therapeutic benefits in animals challenged with hypercaloric dietary regimens. TETA reduced obesity induced by high-fat diet, excessive sucrose intake, or leptin deficiency, as it reduced glucose intolerance and hepatosteatosis, but induced autophagy. Mechanistically, these effects did not involve the depletion of copper from plasma or internal organs. Rather, the TETA effects relied on the activation of an energy-consuming polyamine catabolism, secondary to the stabilization of spermidine/spermine N1-acetyltransferase-1 (SAT1) by TETA, resulting in enhanced enzymatic activity of SAT. All the positive effects of TETA on high-fat diet-induced metabolic syndrome were lost in SAT1-deficient mice. Altogether, these results suggest novel health-promoting effects of TETA that might be taken advantage of for the prevention or treatment of obesity.

Decreased Hepatocyte Autophagy Leads to Synergistic IL-1ß and TNF Mouse Liver Injury and Inflammation.

BACKGROUND AND AIMS: The proinflammatory cytokine IL-1ß has been implicated in the pathophysiology of nonalcoholic and alcoholic steatohepatitis. How IL-1ß promotes liver injury in these diseases is unclear, as no IL-1ß receptor-linked death pathway has been identified. Autophagy functions in hepatocyte resistance to injury and death, and findings of decreased hepatic autophagy in many liver diseases suggest a role for impaired autophagy in disease pathogenesis. Recent findings that autophagy blocks mouse liver injury from lipopolysaccharide led to an examination of autophagy's function in hepatotoxicity from proinflammatory cytokines. APPROACH AND RESULTS: AML12 cells with decreased autophagy from a lentiviral autophagy-related 5 (Atg5) knockdown were resistant to toxicity from TNF, but sensitized to death from IL-1ß, which was markedly amplified by TNF co-treatment. IL-1ß/TNF death was necrosis by trypan blue and propidium iodide positivity, absence of mitochondrial death pathway and caspase activation, and failure of a caspase inhibitor or necrostatin-1s to prevent death. IL-1ß/TNF depleted autophagy-deficient cells of ATP, and ATP depletion and cell death were prevented by supplementation with the energy substrate pyruvate or oleate. Pharmacological inhibitors and genetic knockdown studies demonstrated that IL-1ß/TNF-induced necrosis resulted from lysosomal permeabilization and release of cathepsins B and L in autophagy-deficient cells. Mice with a tamoxifen-inducible, hepatocyte-specific Atg5 knockout were similarly sensitized to cathepsin-dependent hepatocellular injury and death from IL-1ß/TNF in combination, but neither IL-1ß nor TNF alone. Knockout mice had increased hepatic inflammation, and IL-1ß/TNF-treated, autophagy-deficient AML12 cells secreted exosomes with proinflammatory damage-associated molecular patterns. CONCLUSIONS: The findings delineate mechanisms by which decreased hepatocyte autophagy promotes IL-1ß/TNF-induced necrosis from impaired energy homeostasis and lysosomal permeabilization and inflammation through the secretion of exosomal damage-associated molecular patterns.

Biomethane generation from biogas upgrading by means of thin-film composite membrane comprising Linde T and fluorinated polyimide: optimization of fabrication parameters.

Generation of biogas from organic substances is an attractive evolution of energy generation from fossil-based energy supply to renewable resources. In order to exhibit viability in terms of technical execution while being economically feasible, successful purification strategies for biomethane formation must be applicable to industrial gas streams at realistic pressures and temperatures. Membrane-based upgrading technologies have great potential to promote biogas processes because they involve less energy and low maintenance. However, the development of membranes with good polymer-filler contact and minimum defects remains a great challenge. Hitherto, researchers have been making many attempts at developing an established route to fabricate thin-film composite membranes. In the present work, an innovative coupling between Linde T and fluorinated polyimide was employed for biogas upgrading. A facile technique for membrane fabrication was proposed via optimization of the fabrication parameters. The results indicated that composite membrane fabricated with 2 hours of total dispersion duration demonstrated a homogeneous distribution of Linde T particles in the fluorinated polyimide matrix and improved the separation characteristics by up to 172% in upgrading biomethane quality. Thus, the fabricated membrane is feasible to be employed for large-scale and lucrative production with enhanced performance in biogas purification via the feasible fabrication method employed in this work.

Insight into the binding affinity of thiourea in the calcium binding pocket of proteinase K, through high resolution X-ray crystallography.

Proteinase K is a stable serine protease, crystallized and extensively used in the study of molecular interactions at the atomic level. During the current study, crystal structure of proteinase K with thiourea (TU) was solved at 1.45 Å (angstrom) resolution. Proteinase K showed its binding affinity with thiourea after soaking with 200 mM (millimolar) concentration of thiourea solution for 6 h. The binding affinity of proteinase K was evaluated with three different molecules i.e., thiourea, acetamide, and thiosemicarbazide. Interestingly, only the thiourea went into the calcium-binding region, and showed interactions with those amino acids which have also displayed interactions with calcium previously. Pro175 (proline 175), Ser197 (Serine 197), Val198 (valine 198), and Asp200 (aspartic acid 200) were the key amino acids involved in the binding of thiourea with proteinase K. Thiourea showed strong hydrogen bondings with Pro175 (2.85 Å), Ser197 (2.88 Å), and Asp200 (2.90 Å, and 3.30 Å), as the key interactions involved in the binding of thiourea with proteinase K. This study provides an insight into the binding mechanism of thiourea with calcium-binding pocket of proteinase K, and thus can be extrapolated to other calcium-binding proteins.