Alleviation of arthritis through prevention of neutrophil extracellular traps by an orally available inhibitor of protein arginine deiminase 4.

Protein arginine deiminases (PAD) 4 is an enzyme that catalyzes citrullination of protein and its role in autoimmune diseases has been established through clinical genetics and gene knock out studies in mice. Further, studies with PAD4 - deficient mice have shown that PAD4 deficiency does not lead to increased infection or immune suppression, which makes PAD4 an attractive therapeutic target for auto-immune and inflammatory diseases. PAD4 has critical enzymatic role of promoting chromatin decondensation and neutrophil extracellular traps (NETs) formation that is associated with a number of immune-mediated pathological conditions. Here, we present a non-covalent PAD4 inhibitor JBI-589 with high PAD4 isoform selectivity and delineated its binding mode at 2.88 Å resolution by X-ray crystallography. We confirmed its effectiveness in inhibiting NET formation in vitro. Additionally, by using two mouse arthritis models for human rheumatoid arthritis (RA), the well-known disease associated with PAD4 clinically, we established its efficacy in vivo. These results suggest that JBI-589 would be beneficial for both PAD4 and NET-associated pathological conditions.

Genome-wide identification and analysis of the PAL genes from the orchids Apostasia shenzhenica, Dendrobium catenatum and Phalaenopsis equestris.

Phenylalanine ammonia-lyase (PAL) is a key gateway enzyme that connects the phenylpropanoid pathway to primary metabolism. The phenylpropanoid pathway plays a vital role in the growth and environmental adaptation of many plants leading to the production of valuable bioactive compounds with industrial and medical applications. In the present study, nine putative PAL genes from three orchids were identified; five in Apostasia shenzhenica and two each in Dendrobium catenatum and Phalaenopsis equestris. Eighteen motifs and four major conserved functional domains were identified as reported in PAL proteins of other species. All the nine PALs were stable based on their computed physicochemical properties and localized in the cytoplasm. The three-dimensional structures of PALs revealed a homo-tetrameric structure consisting of four identical subunits. A total of 21 cis-regulatory elements with known functions were identified from the promoter regions of all PALs which are responsible for various plant responses to light, stress and growth regulators like auxins, gibberellins and abscisic acid. Phylogenetic analysis showed that the studied PAL proteins clustered in two major clades (clade I and II), placing dicot and monocot PALs in two separate monophyletic clades. In silico gene expression of the identified PALs in different vegetative and reproductive tissues revealed the differential expressions based on tissue type and disclosed that the expression of PAL genes was upregulated in all the tissues examined with an exception of PePAL leaf samples where no expression was detected, however, the same being highly expressed in reproductive tissues (PePAL1-labellum; PePAL2-sepal). In case of AsPALs, the expression was found to be highest in reproductive tissues (AsPAL4-maximum in inflorescence). On the other hand, the expression of DcPALs was found to be highest in vegetative tissues (DcPAL2-maximum in root). Based on the medicinal importance of orchids and the significant role of PAL genes in synthesis of bioactive compounds, the functional characterization of PAL genes can be further exploited in genetic improvement of medicinal orchids.Communicated by Ramaswamy H. Sarma.

ATP citrate lyase inhibitor Bempedoic Acid alleviate long term HFD induced NASH through improvement in glycemic control, reduction of hepatic triglycerides & total cholesterol, modulation of inflammatory & fibrotic genes and improvement in NAS score.

Non-alcoholic fatty liver disease (NAFLD) and Non-alcoholic steatohepatitis (NASH) are chronic liver disorders, the prevalence of which is increasing worldwide. Long term High Fat Diet (HFD) induced NASH animal models closely mimic the characteristics of human NASH and hence used by investigators as a model system for studying the mechanism of action of new drugs. Bempedoic acid (ETC-1002), a ATP citrate lyase (ACLY) inhibitor that lowers the LDL cholesterol was recently approved by US FDA for the treatment of heterozygous familial hypercholesterolemia (HeFH) and established atherosclerotic cardiovascular disease (ASCVD). ACLY is one of the genes modulated in NASH patients and hence we studied the effect of ACLY inhibitor Bempedoic acid in long term HFD induced NASH animal model to understand the pharmacological benefits and the associated mechanism of action of this newly approved drug in NASH. Mice fed with 60% Kcal High Fat Diet for 32 weeks were used for the study and the animals were given Bempedoic acid for 5 weeks at doses of 10 â€‹mg â€‹kg-1, po, qd, and 30 â€‹mg â€‹kg-1, po, qd. Bempedoic acid treatment resulted in inhibition of body weight gain and improved the glycemic control. Bempedoic acid treated group showed statistically significant reduction in plasma ALT, AST, hepatic triglycerides (TG) and total cholesterol (TC), along with statistically significant reduction in steatosis score by histological analysis. Hepatic gene expression analysis showed significant reduction in inflammatory and fibrotic genes such as Mcp-1/Ccl2, Timp-1 & Col1α1. Histological analysis showed significant improvement in NAS score. Overall, Bempedoic acid alleviated HFD induced Non-Alcoholic Steatohepatitis through inhibition of body weight gain, improvement in glycemic control, reduction of hepatic triglycerides & total cholesterol, modulation of inflammatory & fibrotic genes, and improvement in NAS score. Hence, Bempedoic acid can be a potential therapeutic option for metabolic syndrome and NASH.

The complete chloroplast genome of Dendrobium nobile, an endangered medicinal orchid from north-east India and its comparison with related Dendrobium species.

The medicinal orchid genus Dendrobium belonging to the Orchidaceae family is a huge genus comprising about 800-1,500 species. To better illustrate the species status in the genus Dendrobium, a comparative analysis of 33 available chloroplast genomes retrieved from NCBI RefSeq database was compared with that of the first complete chloroplast genome of D. nobile from north-east India based on next-generation sequencing methods (Illumina HiSeq 2500-PE150). Our results provide comparative chloroplast genomic information for taxonomical identification, alignment-free phylogenomic inference and other statistical features of Dendrobium plastomes, which can also provide valuable information on their mutational events and sequence divergence.

In vivo effective dibenzo[b,d]furan-1-yl-thiazoles as novel PDE-4 inhibitors.

Herein we report the synthesis, PDE-4B and TNF-α inhibitory activities of a few dibenzo[b,d]furan-1-yl-thiazole derivatives. The hydroxycyclohexanol amide derivatives 14, 18, 24, 29, 31 and 33 exhibited promising in vitro PDE-4B and TNF-α inhibitory activities. Compound 24 showed good systemic availability in preclinical animal models and was also found to be non-toxic (exploratory mutagenicity test). Further it exhibited promising results in in vivo asthma/COPD and Uveitis models.

A novel animal model of metabolic syndrome with non-alcoholic fatty liver disease and skin inflammation.

CONTEXT: Metabolic syndrome and non-alcoholic fatty liver disease (NAFLD) are the emerging co-morbidities of skin inflammation. Occurrence of skin inflammation such as psoriasis is substantially higher in NAFLD patients than normal. Currently, there are no animal models to study the interaction between these co-morbidities. OBJECTIVE: The present study seeks to develop a simple mouse model of NAFLD-enhanced skin inflammation and to study the effect of NAFLD on different parameters of skin inflammation. MATERIALS AND METHOD: Metabolic syndrome and NAFLD were induced in C57BL/6 mice by feeding high-fat diet (HFD, 60% kcal) and high fructose liquid (HFL, 40% kcal) in drinking water. Skin inflammation was induced by repeated application of oxazolone (1% sensitization and repeated 0.5% challenge) in both normal and NAFLD mice and various parameters of skin inflammation and NAFLD were measured. RESULTS: HFD and HFL diet induced obesity, hyperglycemia, hyperinsulinemia, and histological features of NAFLD in mice. Oxazolone challenge significantly increased ear thickness, ear weight, MPO activity, NF-κB activity, and histological features of skin inflammation in NAFLD mice as compared with normal mice. Overall, induction of oxazolone-induced skin inflammation was more prominent in NAFLD mice than normal mice. Hence, HFD and HFL diet followed by topical oxazolone application develops metabolic syndrome, NAFLD, and enhanced skin inflammation in mice. DISCUSSION AND CONCLUSION: This simple model can be utilized to evaluate a therapeutic strategy for the treatment of metabolic syndrome and NAFLD with skin inflammation and also to understand the nexus between these co-morbidities.

Topical atorvastatin ameliorates 12-O-tetradecanoylphorbol-13-acetate induced skin inflammation by reducing cutaneous cytokine levels and NF-κB activation.

Atorvastatin is a 3-hydroxy-3-methylglutaryl coenzyme-A reductase inhibitor used in the treatment of atherosclerosis and dyslipidemia. Studies have evaluated the utility of statins in the treatment of skin inflammation but with varied results. In the present study, we investigated the effect of atorvastatin on TNF-α release and keratinocyte proliferation in vitro and in acute and chronic 12-O-tetradecanoylphorbol-13-acetate (TPA) induced skin inflammation in vivo. Atorvastatin significantly inhibited lipopolysacharide induced TNF-α release in THP-1 cells and keratinocyte proliferation in HaCaT cells. In an acute study, topical atorvastatin showed dose dependent reduction in TPA induced skin inflammation with highest efficacy observed at 500 µg/ear dose. In chronic study, topical atorvastatin significantly reduced TPA induced ear thickness, ear weight, cutaneous cytokines, MPO activity and improved histopathological features comparable to that of dexamethasone. Atorvastatin also inhibited TPA stimulated NF-κB activation in mouse ear. In conclusion, our results suggest that atorvastatin ameliorates TPA induced skin inflammation in mice at least in part, due to inhibition of cytokine release and NF-κB activation and may be beneficial for the treatment skin inflammation like psoriasis.

Tubastatin, a selective histone deacetylase 6 inhibitor shows anti-inflammatory and anti-rheumatic effects.

Epigenetic modifications represent a promising new approach to modulate cell functions as observed in autoimmune diseases. Emerging evidence suggests the utility of HDAC inhibitors in the treatment of chronic immune and inflammatory disorders. However, class and isoform selective inhibition of HDAC is currently favored as it limits the toxicity that has been observed with pan-HDAC inhibitors. HDAC6, a member of the HDAC family, whose major substrate is α-tubulin, is being increasingly implicated in the pathogenesis of inflammatory disorders. The present study was carried out to study the potential anti-inflammatory and anti-rheumatic effects of HDAC6 selective inhibitor Tubastatin. Tubastatin, a potent human HDAC6 inhibitor with an IC50 of 11 nM showed significant inhibition of TNF-α and IL-6 in LPS stimulated human THP-1 macrophages with an IC50 of 272 nM and 712 nM respectively. Additionally, Tubastatin inhibited nitric oxide (NO) secretion in murine Raw 264.7 macrophages dose dependently with an IC50 of 4.2 µM and induced α-tubulin hyperacetylation corresponding to HDAC6 inhibition in THP-1 cells without affecting the cell viability. Tubastatin showed significant inhibition of paw volume at 30 mg/kg i.p. in a Freund's complete adjuvant (FCA) induced animal model of inflammation. The disease modifying activity of Tubastatin was also evident in collagen induced arthritis DBA1 mouse model at 30 mg/kg i.p. The significant attenuation of clinical scores (~70%) by Tubastatin was confirmed histopathologically and was found comparable to dexamethasone (~90% inhibition of clinical scores). Tubastatin showed significant inhibition of IL-6 in paw tissues of arthritic mice. The present work has demonstrated anti-inflammatory and antirheumatic effects of a selective HDAC6 inhibitor Tubastatin.

Modulation of the cyclooxygenase pathway via inhibition of nitric oxide production contributes to the anti-inflammatory activity of kaempferol.

Kaempferol has been reported to inhibit nitric oxide synthase and cyclooxygenase enzymes in animal models. The present study was designed to investigate whether kaempferol modulates the cyclooxygenase pathway via inhibition of nitric oxide production, which in turn contributes to its anti-inflammatory activity. Investigations were performed using carrageenan induced rat air pouch model. Inflammation was assessed by measurement of nitrites (nitrite, a breakdown product of nitric oxide), prostaglandin-E(2) levels and cellular infiltration in the pouch fluid exudates. To assess the anti-inflammatory effect of the extract, rat air pouch linings were examined histologically. The levels of nitrite and prostaglandin-E(2) in pouch fluid were measured by using Griess assay and ELISA respectively. Cell counts and differential counts were performed using a Coulter counter and Wright-Giemsa stain respectively. Kaempferol when administered orally at 50 and 100mg/kg dose showed significant inhibition of carrageenan induced production of nitrite (40.12 and 59.74%, respectively) and prostaglandin-E(2) generation (64.23 and 78.55%, respectively). Infiltration of the cells into the rat granuloma air pouch was also significantly inhibited by kaempferol. Modulation of cyclooxygenase pathway via inhibition of nitric oxide synthesis significantly contributes to kaempferol's anti-inflammatory activity. The present study characterizes the effects and mechanisms of naturally occurring phenolic flavonoid kaempferol, on inducible nitric oxide synthase expression and nitric oxide production. These results partially explain the pharmacological efficacy of flavonoids in general and kaempferol in particular as anti-inflammatory compounds.

Evaluation of effect of aqueous extract of Enicostemma littorale Blume in streptozotocin-induced type 1 diabetic rats.

The present investigation was undertaken to standardize and study the dose-dependent effect of three weeks treatment with hot and cold aqueous extract of E. littorale (0.5, 1 and 2 g/kg, po) on streptozotocin (STZ) induced type I diabetic (confirmed by histopathology) rats (45 mg/kg, iv single dose). Treatment of rats with STZ produced cardinal signs of diabetes-mellitus like a significant loss of body weight, polyuria and polydipsia. There was also a significant increase in fasting blood glucose levels and AUC(glucose) associated with decrease in insulin levels and AUC(insulin) in STZ-diabetic rats. Treatment with E. littorale hot extract (1 and 2 g/kg) significantly reduced the elevated food intake and water intake, glucose and AUC(glucose) levels of diabetic rats. There was also a significant increase in serum cholesterol, serum triglyceride in the STZ diabetic rats. Treatment with E. littorale hot extract (1 and 2 g/kg) significantly decreased all these elevated levels in diabetic rats. Hot aqueous extract of E. littorale at 0.5 g/kg produced a significant decrease in serum glucose and triglycerides. At this doses serum cholesterol and AUC(glucose) were not found to be altered significantly.TLC finger-print profiles were established for the aqueous extract using HPTLC. Swertiamarin, which was used as a chemical marker, was found to be one of the major components in the hot extract while it was absent in cold extract. The results suggest that E. littorale possesses potential antidiabetic activity and improves lipid profile at a small dose of 0.5 g/kg.

Amelioration of STZ-induced type 1 diabetic nephropathy by aqueous extract of Enicostemma littorale Blume and swertiamarin in rats.

Diabetic nephropathy (DN) is one of the foremost causes of renal failure and a primary cause of diabetes mellitus related death. Previously, we have reported that aqueous extract of Enicostemma littorale has potential antidiabetic activity. In the present study, we have investigated the effect of aqueous extract of E. littorale 1 g/kg, p.o. and swertiamarin 50 mg/kg, p.o. daily for 3 weeks in type 1 DN complications in SD rats. DN was assessed by serum urea, creatinine, lipid profile and water intake levels. Treatment with aqueous extract of E. littorale and swertiamarin significantly decreased serum urea and creatinine and other parameters associated with the development of DN in type 1 diabetic rats. We have also found considerable improvement in histology of glomerular function of aqueous extract of E. littorale and swertiamarin-treated animals.

Anti-diabetic activity of Zingiber officinale in streptozotocin-induced type I diabetic rats.

The fresh and dried rhizome of Zingiber officinale Roscoe (commonly known as ginger) is widely used in traditional medicine. We have studied the effect of the juice of Z. officinale (4 mL kg(-1), p.o. daily) for 6 weeks on streptozotocin (STZ)-induced type I diabetic rats with particular reference to the involvement of serotonin (5-hydroxytryptamine; 5-HT) receptors in glycaemic control. In normoglycaemic rats, 5-HT (1mg kg(-1), i.p.) produced hyperglycaemia and hypoinsulinaemia, which was significantly prevented by the juice of Z. officinale. STZ-diabetes produced a significant increase in fasting glucose levels that was associated with a significant decrease in serum insulin levels. Treatment with Z. officinale produced a significant increase in insulin levels and a decrease in fasting glucose levels in diabetic rats. In an oral glucose tolerance test, treatment with Z. officinale was found to decrease significantly the area under the curve of glucose and to increase the area under the curve of insulin in STZ-diabetic rats. Treatment with Z. officinale also caused a decrease in serum cholesterol, serum triglyceride and blood pressure in diabetic rats. Our data suggest a potential antidiabetic activity of the juice of Z. officinale in type I diabetic rats, possibly involving 5-HT receptors.