Investigation of Protective Effect of Matricaria Chamomilla L. Extract on Methotrexate-Induced Hepatotoxicity in Wistar Rat

Abstract Methotrexate (MTX) was shown to cause oxidative stress and liver damage. The objective was to investigate the possible protective effects of Matricaria Chamomilla L. (chamomile) extract with anti-oxidant and anti-inflammatory properties on the methotrexate-induced liver toxicity. Twenty four Wistar rats were divided into four groups. MTX group was injected intraperitoneally on days 7 and 14 with 20 mg/kg methotrexate. Groups CE200 (chamomile extract 200 mg/kg/day) and CE300 (chamomile extract 300 mg/kg/day) received the same dose of methotrexate added with chamomile extract orally for 15 days at 200 mg/kg and 300 mg/kg respectively and the last group was healthy control group. Results of biochemical analyses indicated serum liver biomarkers (aminotransferases), alkaline phosphatase (ALP), albumin, and liver content of anti-oxidant enzymes (catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px)), reduced glutathione (GSH) and total anti-oxidant capacity (TAC) significantly increased (P <0.05-0.001) to normal in the CE treated groups compared to those of the MTX group. Serum bilirubin and hepatic malondialdehyde (MDA) levels significantly increased (P ˂0.001) in MTX group compared to those of the control group and decreased in CE200 and CE300 groups compared to those of the MTX group. Histopathological study showed inflammatory damage, necrotic cells and lipid infiltration in MTX group. In the groups treated with the chamomile extract, a significant improvement was observed in liver tissue in response to increased dose of the extract. In conclusion, chamomile extract administration could have a protective role in methotrexate-induced liver toxicity in rats through improving anti-oxidant defense system.

Modification of Sexual Hormones in Rheumatoid Arthritis Patients by M2000 (ß-D-mannuronic Acid) as a Novel NSAID with Immunosuppressive Property.

BACKGROUND: Based on in-vitro, in-vivo and human studies, the ß-D-mannuronic acid (M2000) has been introduced as a novel non-steroidal anti-inflammatory drug (NSAID) with immunosuppressive properties. OBJECTIVE: This study aimed to evaluate the efficacy of this drug on serum level of sex hormones (Estradiol, Progesterone, and DHEAS) in rheumatoid arthritis (RA) patients. METHODS: The present research was performed on 10 RA patients who had an inadequate response to conventional treatments (clinical trial identifier: IRCT2014011213739N2). During this trial, the patients were permitted to continue the conventional therapy along with adding M2000 orally at a dose of 500 mg twice daily for 12 weeks. Serum samples were collected in a normal group, patient group (at baseline) and treatment group (after 12 weeks). The samples were tested for evaluating the serum level of Estradiol, Progesterone, and DHEAS using chemiluminescent microparticle immunoassay. RESULTS: Data showed that the serum level of estradiol was reduced (both in men and women) during the treatment with M2000 (after 12 weeks), but there was no significant difference in the non-treated group with M2000 (p > 0.05). In addition, the serum level of progesterone and DHEAS significantly increased following the 12-week administration of M2000 in both male and female patients, compared to the non-treated group with M2000 (p < 0.001, p < 0.05, p < 0.05, p < 0.01, respectively). CONCLUSION: The present research showed that the sex hormones might be modified by M2000 therapy in RA patients by increasing the serum level of progesterone and DHEAS compared to healthy individuals.

Very rapid amyloid fibril formation by a bacterial lipase in the absence of a detectable lag phase.

The conversion of proteins from their soluble states into well-organized amyloid fibrils has received abundant attention. This process typically consists of three stages: lag, growth and plateau phases. In this study, the process of amyloid fibril formation by lipase from Pseudomonas sp. after diluting out urea was examined by Thioflavin T (ThT) fluorescence, Congo red (CR) binding, 8-anilinonaphthalene-1-sulfonic acid (ANS) binding, dynamic light scattering (DLS), circular dichroism (CD) and Fourier transform infrared (FTIR) spectroscopies, X-ray diffraction (XRD) and transmission electron microscopy (TEM). To exclude the presence of preformed aggregates in the pure lipase sample, aforementioned assays were also performed for the protein unfolded in urea before dilution. The aggregates formed immediately after dilution were found to bind to ThT and CR and contain a significant amount of ß-sheet structure, as determined by far-UV CD and FTIR spectroscopies, as well as XRD analysis. Moreover, these aggregates present, at least in part, a fibrillar morphology, as deduced with TEM. This examination showed that lipase fibril formation proceeds quickly after dilution, within a few seconds, without a detectable lag phase. We also investigated bacterial inclusion bodies formed after expression of lipase in E. coli, providing evidence for the existence of rapidly formed amyloid-like structural and tinctorial properties in the lipase-containing inclusion bodies.

Amyloid fibril formation by a normally folded protein in the absence of denaturants and agitation.

The conversion of normally folded proteins into amyloid-like fibrils is an important process in protein chemistry, biology, pathology and biotechnology. This process generally requires harsh conditions, such as pH extremes, organic cosolvents, high temperatures, high pressures or shear forces. Such conditions promote aggregation because they partially unfold structured proteins or allow the sampling of locally unfolded native-like states, both of which possibly represent amyloidogenic states. Here we report the formation of amyloid-like fibrils by the lipase from Pseudomonas sp. under conditions that are close to physiological, that is, in the absence of denaturants and agitation. The resulting aggregates bind thioflavin T and Congo red, causing their characteristic spectral changes observed in the presence of amyloid fibrils. They possess a significant quantity of ß-sheet structure, as detected with Fourier transform infrared and far-UV circular dichroism spectroscopies, and appear fibrillar using transmission electron microscopy. These results indicate that the lipase from Pseudomonas sp. can be a useful model system for the characterization of a key process, such as amyloid fibril formation under physiological conditions.

Comparison of the molten globule states of thermophilic and mesophilic alpha-amylases.

In recent years great interest has been generated in the process of protein folding, and the formation of intermediates during the folding process has been proven with new experimental strategies. In the present work, we have examined the molten globule state of Bacillus licheniformis alpha-amylase (BLA) by intrinsic fluorescence and circular dichroism spectra, 1-anilino naphthalene-8-sulfonate (ANS) binding and proteolytic digestion by pepsin, for comparison to its mesophilic counterpart, Bacillus amyloliquefaciens alpha-amylase (BAA). At pH 4.0, both enzymes acquire partially folded state which show characteristics of molten globule state. They unfold in such a way that their hydrophobic surfaces are exposed to a greater extent compared to the native forms. Chemical denaturation studies by guanidine hydrochloride and proteolytic digestion with pepsin show that molten globule state of BLA is more stable than from BAA. Results from gel filtration indicate that BAA has the same compactness at pH 4.0 and 7.5. However, molten globule state of BLA is less compact than its native state. The effects of polyols such as trehalose, sorbitol and glycerol on refolding of enzymes from molten globule to native state were also studied. These polyols are effective on refolding of mesophilic alpha-amylase but only slightly effect on BLA refolding. In addition, the folding pathway and stability of intermediate state of the thermophilic and the mesophilic alpha-amylases are discussed.