Abu Asad Chowdhury

Endogenous 15-deoxy-Δ12,14-prostaglandin J2 synthesized by adipocytes during maturation phase contributes to upregulation of fat storage

15‐Deoxy‐Δ12,14‐prostaglandin J2 (15d‐PGJ2) has been identified as a natural ligand for peroxisome proliferator‐activated receptor (PPAR) γ to promote adipogenesis. However, it remains elusive about the ability of PPARγ‐expressing adipocytes to produce PGJ2 series and the role in the life cycle of adipocytes. Here, we developed an enzyme‐linked immunosorbent assay specific for 15d‐PGJ2. The analysis using this method revealed the increase in the endogenous synthesis of immunoreactive 15d‐PGJ2 in cultured adipocytes during the maturation phase. Further studies using cyclooxygenase inhibitors clarified the contribution of endogeous 15d‐PGJ2 produced by mature adipocytes to upregulation of fat storage in an autocrine manner.

Development of enzyme-linked immunosorbent assay for Δ12-prostaglandin J2 and its application to the measurement of the endogenous product generated by cultured adipocytes during the maturation phase.

Peroxisome proliferator-activated receptor (PPAR)γ is a well-known master regulator for the differentiation and maturation of adipocytes. Prostaglandin (PG) D(2) can be produced in adipocytes and dehydrated to J(2) series of PGs including 15-deoxy-Δ(12,14)-PGJ(2) (15d-PGJ(2)) and Δ(12)-PGJ(2), which serve as pro-adipogenic prostanoids through the activation of PPARγ. However, the quantitative determination of Δ(12)-PGJ(2) has not been attempted during the life stage of adipocytes. In this study, we developed an enzyme-linked immunosorbent assay using mouse antiserum specific for Δ(12)-PGJ(2). According to the standard curve, the amount of Δ(12)-PGJ(2) can be measured from 0.5 pg to 14.4 ng in an assay. Our antiserum did not recognize most other prostanoids including 15d-PGJ(2), while it only showed the cross-reaction of 28% with unstable PGJ(2). This immunological assay was applied to the determination of the endogenous formation of Δ(12)-PGJ(2) in cultured 3T3-L1 adipocytes during the maturation phase. The ability of cultured adipocytes to form endogenous Δ(12)-PGJ(2) increased gradually at an earlier stage of the maturation phase and detectable at higher levels than 15d-PGJ(2). Treatment of cultured cells with either aspirin or indomethacin, a general cyclooxygenase inhibitor, significantly reduced the production of endogenous Δ(12)-PGJ(2) in the maturation medium as expected. Furthermore, we evaluated individually the exogenous effects of PGJ(2) series at various doses on adipogenesis during the maturation phase. Although Δ(12)-PGJ(2) was slightly less potent than 15d-PGJ(2), each of these PGJ(2) series rescued effectively both the accumulation of fats and the gene expression of typical adipocyte-markers that were attenuated in the presence of aspirin. Taken together, our findings indicate that endogenous Δ(12)-PGJ(2) contributes substantially to the up-regulation of adipogenesis program through the activation of PPARγ together with 15d-PGJ(2) during the maturation phase of cultured adipocytes.

Chemical and biological studies of Kalanchoe pinnata (Lam.) growing in Bangladesh

Abstract Objective To isolate compounds from K. pinnata and elucidate their structures and to explore preliminary antioxidant, antimicrobial, cytotoxic and thombolytic activities of extractives of the plant. Methods The methanol extract of whole plant of K. pinnata has been subjected to different chromatographic separation and purification processes to isolate the secondary metabolites. The structures of the isolated compounds have been elucidated by extensive NMR studies. The free radical scavenging activity of the crude extract and its different Kupchan fractions were determined on stable radical DPPH. In vitro antimicrobial activity was determined by the disk diffusion method. Cytotoxicity screening has been performed against Artemia salina . Total phenolics content, membrane stabilizing activity and thombolytic activities were assessed by following established protocol. Results The isolated compounds were identified as glut-5(6)-en-3-one, taraxerone, 3β-friedelanol, β-amyrin-3-acetate, 3,5,7,3′,5′-pentahydroxyflavone and β-sitosterol. The chloroform soluble fraction showed potent antioxidant activity of (IC 50 = 80.0 μg/mL) and significant cytotoxicity, while the crude extract demonstrated noticeable total polyphenol content (149.24 mg of GAE/gm of extractive), moderate membrane stabilizing activity and inhibition of clot lysis of blood. Conclusions The obtained results rationalize the folkloric use of the plant and can be further investigated to isolate the active compounds responsible for the biological activities.

Stable expression of lipocalin-type prostaglandin D synthase in cultured preadipocytes impairs adipogenesis program independently of endogenous prostanoids.

Lipocalin-type prostaglandin D synthase (L-PGDS) expressed preferentially in adipocytes is responsible for the synthesis of PGD(2) and its non-enzymatic dehydration products, PGJ(2) series, serving as pro-adipogenic factors. However, the role of L-PGDS in the regulation of adipogenesis is complex because of the occurrence of several derivatives from PGD(2) and their distinct receptor subtypes as well as other functions such as a transporter of lipophilic molecules. To manipulate the expression levels of L-PGDS in cultured adipocytes, cultured preadipogenic 3T3-L1 cells were transfected stably with a mammalian expression vector having cDNA encoding murine L-PGDS oriented in the sense direction. The isolated cloned stable transfectants with L-PGDS expressed higher levels of the transcript and protein levels of L-PGDS, and synthesized PGD(2) from exogenous arachidonic acid at significantly higher levels. By contrast, the synthesis of PGE(2) remained unchanged, indicating no influence on the reactions of cyclooxygenase (COX) and PGE synthase. Furthermore, the ability of those transfectants to synthesize Δ(12)-PGJ(2) increased more greatly during the maturation phase. The sustained expression of L-PGDS in cultured stable transfectants hampered the storage of fats during the maturation phase of adipocytes, which was accompanied by the reduced gene expression of adipocyte-specific markers reflecting the down-regulation of the adipogenesis program. The suppressed adipogenesis was not rescued by either exogenous aspirin or peroxisome proliferator-activated receptor γ (PPARγ) agonists including troglitazone and Δ(12)-PGJ(2). Taken together, the results indicate the negative regulation of the adipogenesis program by the enhanced expression of L-PGDS through a cellular mechanism involving the interference of the PPARγ signaling pathway without the contribution of endogenous pro-adipogenic prostanoids.

15-Deoxy-Δ12,14-prostaglandin J2 interferes inducible synthesis of prostaglandins E2 and F2α that suppress subsequent adipogenesis program in cultured preadipocytes

Cultured preadipocytes enhance the synthesis of prostaglandin (PG) E(2) and PGF(2α) involving the induction of cyclooxygenase (COX)-2 during the growth phase upon stimulation with a mixture of phorbol 12-myristate 13-acetate, a mitogenic factor, and calcium ionophore A23187. Here, we studied the interactive effect of 15-deoxy-Δ(12,14)-prostaglandin J(2) (15d-PGJ(2)) on the inducible synthesis of the endogenous PGs in cultured preadipocytes and its implication in adipogenesis program. 15d-PGJ(2) interfered significantly the endogenous synthesis of those PGs in response to cell stimuli by suppressing the induction of COX-2 following the attenuation of NF-κB activation. In contrast, Δ(12)-PGJ(2) and troglitazone had almost no inhibitory effects, indicating a mechanism independent of the activation of peroxisome proliferator-activated receptor γ for the action of 15-PGJ(2). Pyrrolidinedithiocarbamate (PDTC), an NF-κB inhibitor, effectively inhibited on the inducible synthesis of those PGs in preadipocytes. Endogenous PGs generated by preadipocytes only during the growth phase in response to the cell stimuli autonomously attenuated the subsequent adipogenesis program leading to the differentiation and maturation of adipocytes. These effects were prevented by additional co-incubation of preadipocytes with either 15d-PGJ(2) or PDTC although 15d-PGJ(2) alone has no stimulatory effect. Moreover, 15d-PGJ(2) did not block the inhibitory effects of exogenous PGE(2) and PGF(2α) on the adipogenesis program in preadipocytes. Taken together, 15d-PGJ(2) can interfere the COX pathway leading to the induced synthesis of endogenous PGs that contribute to negative regulation of adipogenesis program in preadipocytes.

Sustained expression of lipocalin-type prostaglandin D synthase in the antisense direction positively regulates adipogenesis in cloned cultured preadipocytes.

Adipocytes express preferentially lipocalin-type prostaglandin (PG)D synthase (L-PGDS) that is responsible for the biosynthesis of PGD(2) and other related prostanoids with pro-adipogenic or anti-adipogenic effects. To evaluate the role of L-PGDS in cultured adipocytes and the precursor cells, we attempted to interfere the intracellular expression of L-PGDS in cultured 3T3-L1 preadipocytes by stable transfection with a mammalian expression vector having the full-length cDNA of L-PGDS oriented in the antisense direction. The cloned transfectants with antisense L-PGDS exhibited the reduction in the transcript and protein levels of L-PGDS, resulting in the significant inhibition of the PGD(2) synthesis from exogenous and endogenous arachidonic acid. By contrast, the synthesis of PGE(2) was not influenced appreciably, indicating no interfering effects on cyclooxygenases and PGE synthases. The stable transfection with antisense L-PGDS induced markedly the stimulation of fat storage in cultured adipocytes during the maturation phase. In addition, the spontaneous accumulation of fats occurred in the transfectants with antisense L-PGDS without undergoing the stimulation with inducing factors. The gene expression studies revealed the enhanced expression of adipocyte-specific markers in the transfectants with antisense L-PGDS, indicating the up-regulation of adipogenesis program. The stimulated adipogenesis was significantly reversed by anti-adipogenic prostanoids including PGE(2) and PGF(2α), while the storage of fats was additionally enhanced by pro-adipogenic 15-deoxy- Δ(12,14)-prostaglandin J(2). These results suggest that the stably reduced expression levels of L-PGDS regulates positively adipogenesis program in a cellular mechanism independent of pro-adipogenic action of PGJ(2) series.

Microbial production of biogas form organic wastes

Abstract Biogas production through bacterial digestion of water hyacinth, rice straw and sawdust inoculated with cow dung was investigated in a batch loading of laboratory digestors. It was found that the overall gas yield was 368.00 l from water hyacinth having 77.13% methane, 310.00 l from rice straw with 77.80% methane, and 144.80 l from sandust containing 62.28% methane. Along with the production of gas, the change in pH and microbial population were also monitored. The pH of the digestion mixture was initially low and found to be increasing as the digestion progressed. The total microbial population of bacteria, fungi, actinomycetes, cellulolytic organisms, and coliform bacteria were found to decrease with time.

A Study of Prophylactic Effect Against Diabetes of Two Ayurvedic Drugs ‘Jambadyarista’ and ‘Bohumutrantak Ras’ in Normal as well as Alloxan-induced Diabetic Rats

Aims: The present study was designed to evaluate the anti -diabetic as well as prophylactic activity of Jambadyarista and Bohumutrantak Ras in alloxan -induced diabetic rats. Study Design:Study the prophylactic and antiglycemic effects against diabetes of two

A Monoclonal Antibody Specific for Δ12-Prostaglandin J2 and Its Utilization in the Immunological Assay in Cell Culture System of Adipocytes

Prostaglandin (PG) D(2) can be produced in adipocytes and dehydrated to PGs of J(2) series, including Δ(12)-PGJ(2) and 15-deoxy-Δ(12,14)-PGJ(2) (15d-PGJ(2)), which serve as pro-adipogenic prostanoids through the activation of peroxisome proliferator-activated receptor γ. To accomplish the quantification of Δ(12)-PGJ(2) in the cell culture system of adipocytes, the present study aimed to develop a sensitive and specific immunological assay for Δ(12)-PGJ(2). Here, we established a cloned hybridoma cell line secreting a monoclonal antibody specifically recognizing Δ(12)-PGJ(2) and utilized for the development of its solid-phase enzyme-linked immunosorbent assay (ELISA). The immobilized antigen using a conjugate of Δ(12)-PGJ(2) and γ-globulin was competitively allowed to react with the monoclonal antibody in the presence of free Δ(12)-PGJ(2). The assay provided a sensitive calibration curve for Δ(12)-PGJ(2), allowing us to determine a range from 0.16 pg to 0.99 ng with a value of 13 pg at 50% displacement in one assay. The monoclonal antibody showed almost no cross-reactivity with other related prostanoids since PGJ(2) and 15d-PGJ(2) were only recognized with much lower values of 0.5% and 0.2%, respectively. The accuracy for determining Δ(12)-PGJ(2) in the culture medium of adipocytes was confirmed by measurement after the culture medium was fortified with known amounts of authentic Δ(12)-PGJ(2) in a range from 10 to 200 pg/mL. The application of our ELISA revealed that the formation of Δ(12)-PGJ(2) became more pronounced after several hours of incubation of PGD(2) at 37°C in fresh maturation medium of cultured adipocytes. Furthermore, we provide evidence for the increased ability of cultured adipocytes to synthesize endogenous Δ(12)-PGJ(2) during the progression of adipogenesis. These results indicate the reliability and usefulness of our solid-phase ELISA for stable Δ(12)-PGJ(2), reflecting the biosynthesis of unstable PGD(2) in the culture system of adipocytes.

Cardiovascular Activities of an Ayurvedic Preparation Amalaki Rasayan in Rat Model

The importance of traditional or alternative medicines including Ayurvedic drugs are increasing day by day because of their necessary pharmacological actions and fewer side effects. Previous investigators showed hypolipidemic and cardioprotective effects of Amla which is the active ingredient of Amalaki Rasayan. It is an ayurvedic formulation that contains extracts of Phyllanthus emblica plant. Therefore, this study has been conducted to investigate the scientific evidence of its cardioprotective effects in rats by recording electrocardiographic tracings. Rats, of either sex, were treated with preparation containing extracts of Phyllanthus emblica in doses 800 μg/kg bodyweight, 1600 μg/kg body weight and 3200 μg/kg body weight through intraperitoneal route. Electrocardiogram along with heart rate and other parameters were measuredin rat model before and after administration of Amalaki Rasayan. It was evident that Amalaki Rasayan, at a dose of 800 μg/kg, was safe but 1600 μg/kg and 3200 μg/kg produced abnormal activities in the heart. It can be inferred from the study that, the ayurvedic preparation Amalaki Rasayan possesses significant cardioprotective effect.