Arjunolic acid, a peroxisome proliferator-activated receptor α agonist, regresses cardiac fibrosis by inhibiting non-canonical TGF-ß signaling.

Cardiac hypertrophy and associated heart fibrosis remain a major cause of death worldwide. Phytochemicals have gained attention as alternative therapeutics for managing cardiovascular diseases. These include the extract from the plant Terminalia arjuna, which is a popular cardioprotectant and may prevent or slow progression of pathological hypertrophy to heart failure. Here, we investigated the mode of action of a principal bioactive T. arjuna compound, arjunolic acid (AA), in ameliorating hemodynamic load-induced cardiac fibrosis and identified its intracellular target. Our data revealed that AA significantly represses collagen expression and improves cardiac function during hypertrophy. We found that AA binds to and stabilizes the ligand-binding domain of peroxisome proliferator-activated receptor α (PPARα) and increases its expression during cardiac hypertrophy. PPARα knockdown during AA treatment in hypertrophy samples, including angiotensin II-treated adult cardiac fibroblasts and renal artery-ligated rat heart, suggests that AA-driven cardioprotection primarily arises from PPARα agonism. Moreover, AA-induced PPARα up-regulation leads to repression of TGF-ß signaling, specifically by inhibiting TGF-ß-activated kinase1 (TAK1) phosphorylation. We observed that PPARα directly interacts with TAK1, predominantly via PPARα N-terminal transactivation domain (AF-1) thereby masking the TAK1 kinase domain. The AA-induced PPARα-bound TAK1 level thereby shows inverse correlation with the phosphorylation level of TAK1 and subsequent reduction in p38 MAPK and NF-κBp65 activation, ultimately culminating in amelioration of excess collagen synthesis in cardiac hypertrophy. In conclusion, our findings unravel the mechanism of AA action in regressing hypertrophy-associated cardiac fibrosis by assigning a role of AA as a PPARα agonist that inactivates non-canonical TGF-ß signaling.

Characterization of lignin-degrading enzymes (LDEs) from a dimorphic novel fungus and identification of products of enzymatic breakdown of lignin.

Lignin is a major component of all plants, the degradation of which remains a major challenge to date owing to its recalcitrant nature. Several classes of fungi have been studied to carry out this process to some extent, but overall the process remains inefficient. We have isolated a novel alkalophilic dimorphic lignin-degrading Deuteromycete from soil, identified as "uncultured" and coded as MVI.2011. Supernatant from 12-h culture of MVI.2011 in optimized mineral medium containing lignin pH 9.0 was analysed for Lignin Peroxidase, Manganese Peroxidase and Laccase. Enzyme purification was carried out by standard protocols using ammonium sulphate precipitation followed by further purification by Gel Permeation Chromatography. Analysis of total protein, specific enzyme activity and molecular weight of the GPC-purified LiP, MnP and Laccase showed 93.83 µg/ml, 5.27 U/mg, 42 kDa; 78.13 µg/ml, 13.18 U/mg, 45 kDa and 85.81 µg/ml, 4.77 U/mg, 62 kDa, respectively. The purified enzymes possessed high activity over a wide range of pH (4-11), and temperature (30-55 °C). The optimum substrate concentration was 20 µg/ml of lignin for all the three enzymes. CD spectra suggested that the predominant secondary structure was helix in LiP, and, turns in MnP and Laccase. The breakdown products of lignin degradation by MVI.2011 and the three purified enzymes were detected and identified by FTIR and GC-MS. They were oxalic acid, hentriacontane, derivatives of octadecane, nonane, etc. These vital compounds are certain to find application as biofuels, an alternate energy source in various industries.

Isolation, characterisation and cytotoxicity study of arjunolic acid from Terminalia arjuna.

Arjunolic acid (AA), a triterpenoid, was isolated from the ethyl acetate and methanol extracts of Terminalia arjuna core wood. The purity of AA was analysed by its melting point, FT-IR and NMR spectroscopy analyses. In vitro cytotoxicity was assessed using Ehrlich ascites carcinoma (EAC) and Dalton's lymphoma (DAL) cell lines by incubating with different concentrations of AA. The cancer cell death percentage at 100 µg concentrations of AA ranged between 66% and 70% on the DAL and EAC cell lines, respectively. This infers that AA causes considerable membrane damage to cancer cells.

Papilloma virus in cervical carcinoma: detection of viral antigen in cancer cells.

Paraffin sections of biopsies from histopathologically confirmed cases of uterine cervical carcinoma, cervical scrapings from dysplasia, chronic cervicitis, tumour cells from carcinoma of the oral cavity and normal tissues from healthy normal cervix and oral cavity scrapings were examined for the presence of Human papilloma virus antigens. The techniques adopted were the Indirect Immunofluorescence Stainig and the Peroxide-Anti-Peroxidase techniques. The HPV-antigen was present in 38 percent and 41 percent of invasive carcinoma cervix, by PAP and IIF methods respectively. In cervical dysplasia 8-13% revealed HPV antigen while oral carcinoma cells and normal tissue samples were totally negative.

Detection of herpes simplex virus type-2 DNA and human papilloma virus DNA sequences in cervical carcinoma tissue by molecular hybridization.

Biopsy samples from one hundred and two patients with squamous cell carcinoma of the uterine cervix and tissues from twelve healthy normal cervical tissues (post hysterectomy) were examined for HSV-type 2 and HPV-11 DNA sequences by molecular hybridization technique. In the carcinoma tissue extracts 53% contained HSV-2 DNA, 27% -HSV-1-DNA and 36% showed HPV-11 gene sequences while 5.7% were found to contain both HPV and HSV-2 DNA. Biopsies from healthy cervix were completely negative for HSV-2 and HPV-11 DNA sequences.

Detection of herpes simplex virus type-2 antigen(s) in biopsies from carcinoma of the uterine cervix.

Indirect immunofluorescence technique was employed to detect herpes simplex virus type-2 (HSV-2) antigens in tumor biopsies from 215 patients with carcinoma of the uterine cervix. A total of 169 samples (79%) revealed brilliant nuclear fluorescence. Inflammatory cells infiltrating the tumor mass were positive to 60 of the 215 patients (28%). Samples showed no significant variation in the degree of fluorescence or proportion of cells binding HSV antibody with advancement in the clinical stage of the disease. Fluorescence was totally abolished when incubated with HSV-2 antiserum absorbed with a specific homologous virus. Among controls, there was fluorescence in 27% of cervical scrapings from normal women and 34% (42/124) among patients with gynecological disorders other than cervical malignancy. In cervical dysplasia 23 out of 40 patients (58%) expressed herpes virus-associated antigens. There was membrane fluorescence in live malignant cell preparations in 3 of 28 patients (11%). Normal cervix tissue from hysterectomy specimens and breast cancer cells were negative for herpes simplex virus-related antigens. Pre-immune serum and PBS showed nonspecific fluorescence in 25% and 23% of sera, respectively.