In vivo investigation of the anti-liver fibrosis impact of Balanites aegyptiaca/ chitosan nanoparticles.

Balanites aegyptiaca (B. aegyptiaca) is an African herb with traditional medical applications. Various pathogenic factors cause hepatic fibrosis and require novel treatment alternatives. Nanoformulation-based natural products can overcome the available drug problems by increasing the efficacy of natural products targeting disease markers. The current study investigated B. aegyptiaca methanolic extract using high-pressure liquid chromatography (HPLC), and B. aegyptiaca/chitosan nanoparticles were prepared. In vivo, evaluation tests were performed to assess the curative effect of the successfully prepared B. aegyptiaca/chitosan nanoparticles. For 30 days, the rats were divided into six groups, typical and fibrosis groups, where the liver fibrosis groups received B. aegyptiaca extract, silymarin, chitosan nanoparticles, and B. aegyptiaca/chitosan nanoparticles daily. In the current investigation, phenolic molecules are the major compounds detected in B. aegyptiaca extract. UV showed that the prepared B. aegyptiaca /chitosan nanoparticles had a single peak at 280 nm, a particle size of 35.0 ± 6.0 nm, and a negative charge at - 8.3 mV. The animal studies showed that the synthetic B. aegyptiaca/chitosan nanoparticles showed substantial anti-fibrotic protective effects against CCl4-induced hepatic fibrosis in rats when compared with other groups through optimization of biochemical and oxidative markers, improved histological changes, and modulated the expression of Col1a1, Acta2 and Cxcl9 genes, which manage liver fibrosis. In conclusion, the current research indicated that the prepared B. aegyptiaca/chitosan nanoparticles improved histological structure and significantly enhanced the biochemical and genetic markers of liver fibrosis in an animal model.

Aspergillus Niger thermostable Cytosine deaminase-dextran conjugates with enhanced structure stability, proteolytic resistance, and Antiproliferative activity.

Cytosine deaminase (CDA) is a prodrug mediating enzyme converting 5-flurocytosine into 5-flurouracil with profound broad-range anticancer activity towards various cell lines. Availability, molecular stability, and catalytic efficiency are the main limiting factors halting the clinical applications of this enzyme on prodrug and gene therapies, thus, screening for CDA with unique biochemical and catalytic properties was the objective. Thermotolerant/ thermophilic fungi could be a distinctive repertoire for enzymes with affordable stability and catalytic efficiency. Among the recovered thermotolerant isolates, Aspergillus niger with optimal growth at 45 °C had the highest CDA productivity. The enzyme was purified, with purification 15.4 folds, molecular mass 48 kDa and 98 kDa, under denaturing and native PAGE, respectively. The purified CDA was covalently conjugated with dextran with the highest immobilization yield of 75%. The free and CDA-dextran conjugates have the same optimum pH 7.4, reaction temperature 37 °C, and pI 4.5, and similar response to the inhibitors and amino acids suicide analogues, ensuring the lack of effect of dextran conjugation on the CDA conformational structure. CDA-Dextran conjugates had more resistance to proteolysis in response to proteinase K and trypsin by 2.9 and 1.5 folds, respectively. CDA-Dextran conjugates displayed a dramatic structural and thermal stability than the free enzyme, authenticating the acquired structural and catalytic stability upon dextran conjugation. The thermal stability of CDA was increased by about 1.5 folds, upon dextran conjugation, as revealed from the half-life time (T1/2). The affinity of CDA-conjugates (Km 0.15 mM) and free CDA (Km 0.22 mM) to deaminate 5-fluorocytosine was increased by 1.5 folds. Upon dextran conjugation, the antiproliferative activity of the CDA towards the different cell lines "MDA-MB, HepG-2, and PC-3" was significantly increased by mediating the prodrug 5-FC. The CDA-dextran conjugates strongly reduce the tumor size and weight of the Ehrlich cells (EAC), dramatically increase the titers of Caspase-independent apoptotic markers PARP-1 and AIF, with no cellular cytotoxic activity, as revealed from the hematological and biochemical parameters.

Barhi date (Phoenix dactylifera) extract ameliorates hepatocellular carcinoma in male rats.

Hepatocellular carcinoma (HCC) is a malignant tumor with limited treatment options. Given this fact, it may be important to develop new molecular targeted therapies from natural products, especially those which are primary sources of effective anticancer drugs with distinct mechanisms. Moreover, the complementary use of traditional herbs or fruit may increase the possibility of finding curative options for cancer. Here we explore the anticancer effects and possible molecular mechanism of Barhi date extract using an HCC rat model. Thirty two male albino rats were arbitrarily allocated into four groups: a negative control group (NCG); a positive control group (PCG), which received CCl4 (1 ml/kg b.wt./ i.p.) twice a week for three months; a Barhi date extract (400 mg/kg b.wt./day/orally) treatment group (DTG) during the third month of CCl4 administration; and a cisplatin (1.5 mg/kg b.wt./ i.p.) treatment group ( CTG) during the third month of CCl4 administration. After treatment we performed biochemical analyses of all groups to assess relative eukaryotic initiation factor 2 alpha (eIF2α), extracellular signal-regulated kinases (ERKs), protein kinase RNA-like endoplasmic reticulum kinase (PERK), poly (ADP-ribose) polymerase (PARP), and CASPASE 3 protein content, and examined expression of the genes phosphatase and tensin homolog (PTEN) and protein kinase B (AKT). We also performed an immunohistochemistry assay for alpha-fetoprotein (AFP). Our data showed higher PARP and CASPASE3 levels and liver enzymes (alanine aminotransferase [ALT], aspartate aminotransferase [AST], and alkaline phosphatase [ALP]) in the PCG compared to the DTG and the cisplatin treatment group CTG. However, we also found a significant decrease in PTEN in the PCG relative to both the DTG and the CTG. We conclude that the anti-tumor activity of Barhi date extract may be mediated by the inhibition of cell proliferation and apoptosis via the ERK /PARP/caspase3 pathway and the AKT/ PTEN signaling pathways.

Oral supplementation of policosanol alleviates carbon tetrachloride-induced liver fibrosis in rats.

Liver fibrosis is a prevalent liver disease that requires rapid and effective treatment prior to its progression to cirrhosis and liver damage. Recently, several reports have investigated the efficacy of phytotherapy using natural herbal extracts rather than synthetic drugs to treat several liver diseases. Policosanol is a herbal extract used to treat patients with cardiovascular. However, its therapeutic effect on liver fibrosis is still unknown. Therefore, the present study aimed to assess the potential antifibrotic effect of policosanol compared to silymarin and the possible underlying molecular mechanisms. Rats were categorized into four groups; negative control group "NCG", the fibrotic group "FG", silymarin treated group "STG", and policosanol treated group "PTG". Serum liver enzymes, oxidative stress markers, angiogenic growth factors, and pro-inflammatory cytokines were measured biochemically. The relative mRNA expressions of liver caspase-3 and alpha-smooth muscle actin (α-SMA) were assessed. Immunohistochemical staining was carried out using anti- α-SMA, and anti-caspase-3 antibodies. Compared to NCG, the FG group demonstrated a significant decrease in the level of serum liver enzymes "GSH, TAC, and SDF. Nevertheless, it demonstrateda significant increase in the level of pro-inflammatory cytokines "Il-6, TNF"; oxidative stress markers "NO, MDA", and angiogenic growth factors "VEGF and PDGF" and the expression of α-SMA, and Caspase-3. Interestingly, the values of these measurements were restored to normal levels in the treated groups, particularly the PTG. In conclusion, our data revealed the beneficial effects of co-administration of policosanol or silymarin on the fibrotic liver rat model and thus could be a promising natural therapeutic drug.

Azelastine a potent antihistamine agent, as hypolipidemic and modulator for aortic calcification in diabetic hyperlipidemic rats model.

AIM: Our study aimed to illustrate the effect of the antihistaminic drug azelastine on aortic calcification in diabetic hyperlipidemic (DH) rats along with the underlying molecular mechanism. METHODS: Twenty-four male albino Wistar rats were categorised into four groups. One group received normal rodent chow (normal group), while the other groups were rendered diabetic and hyperlipidemic; one received no drugs and served as a positive control while the other two groups received either azelastine (4 mg/kg) or 10-dehydrogingerdione (10 mg/kg) orally and daily for 8 weeks. RESULTS: Azelastine significantly reduced blood glucose, HbA1c and serum ALP, OCN, downregulated apo B, improved the lipid profile (LDL-c decrease and HDL-c increase), attenuated calcium deposition and aortic calcification as compared to control group. 10-DHGD showed comparatively lower effect. CONCLUSION: Anti-calcifying effect of azelastine might be related to upregulation of apo A (HDL-c) and downregulation of apo B mRNA expression indeed good modulator of aortic calcification. IMPACT STATEMENT: Many studies have indicated that high-density lipoprotein-cholesterol (HDL-c) is inversely correlated with atherosclerotic plaque progression and could reduce cardiovascular disease risk. An anti-calcifying effect of HDL-c has been reported and targeting this lipoprotein may therefore be a valuable approach to vascular calcification control. Azelastine is a selective H1 antagonist that was identified to increase mRNA expression of apolipoprotein A. This encouraged us to investigate the effect of azelastine on lipid profile and markers of aortic calcification in DH rats. Our findings showed that azelastine ameliorated aortic calcification and increased apoA expression along with a decline in apo B. This may represent the underlying mechanism while the histopathological findings offered a significant support to the collected biochemical data.

Bee venom ameliorates cardiac dysfunction in diabetic hyperlipidemic rats.

High levels of blood glucose and lipids are well-known risk factors for heart diseases. Bee venom is a natural product that has a potent hypoglycemic, hypolipidemic, anti-inflammatory, and antioxidant effects. The current study aimed to determine the bee venom effects on cardiac dysfunction compared to combined therapy of metformin and atorvastatin in diabetic hyperlipidemic rats. The median lethal dose of bee venom was estimated, and then 50 adult male albino rats were categorized into five groups. One group was fed a standard diet and served as a negative control, while the other groups were given nicotinamide and streptozotocin injections to induce type 2 diabetes. After confirming diabetes, the rats were fed a high-fat diet for four weeks. The four groups were divided as follows: one group served as a positive control, whereas the other three groups were treated with bee venom (0.5 mg/kg), bee venom (1.23 mg/kg), and combined therapy of metformin (60 mg/kg) and atorvastatin (10 mg/kg), respectively, for four weeks. Upon termination of the experiment, blood samples and heart tissue were obtained. Administration of bee venom using both doses (0.5 and 1.23 mg/kg) and combined therapy of metformin and atorvastatin revealed a significant decrease in the concentrations of glucose, total cholesterol, triacylglycerol, low-density lipoprotein cholesterol, very low-density lipoprotein cholesterol, troponin I, creatine kinase, and lactate dehydrogenase activities. Moreover, a significant decrease had been detedcted in malondialdehyde, nuclear factor-kappa-ß levels, and relative mRNA expression of vascular cell adhesion molecule-1 and galectin-3 in heart tissue compared to the positive control (P < 0.0001). Furthermore, there was a significant increase in bodyweight levels of insulin, high-density lipoprotein cholesterol, and total antioxidant capacity in heart tissue compared to the positive control (P < 0.0001). The results indicate that bee venom can ameliorate cardiac dysfunction through attenuating oxidative stress and downregulating the NF-κß signaling pathway.

Losartan and azelastine either alone or in combination as modulators for endothelial dysfunction and platelets activation in diabetic hyperlipidemic rats.

AIM: The present study aimed mainly to demonstrate the effect of the antihistamine azelastine (AZ) and Angiotensin receptor blocker ( ARB), represented by losartan (LOS) either alone or in combined form on certain metabolic aspects, endothelial dysfunction and platelets activation markers in diabetic hyperlipidemic rat model. METHODS: Rats were randomly classified to five groups: One group fed normal chow diet (NC). Four groups received alloxan and CCT-diet. One group received no treatment (DHC while the other three groups received AZ, LOS and their combination form, respectively for 8 weeks. Serum and tissue samples were collected for biochemical and histological evaluations. RESULTS: DHC rats demonstrated significant hyperglycaemia, dyslipidemia, disturbances in endothelial and platelet activation markers. AZ or LOS administration demonstrated hypoglycaemic and hypolipidemic effects. VCAM-1 and sE-selectin (Endothelial function markers) along with CD63 (Platelet activation marker) showed significant decrease as compared to control group. AZ administration exerted little prominent effects than that of LOS, while their combination demonstrated remarkable changes compared to monotherapy. Histopathological findings were in agreement to certain extent with the biomarkers results. CONCLUSIONS: Both drug categories may be expressed as suitable therapeutic tools for atherosclerotic complications either alone or along with other hypolipidemic drugs.

Exploiting the Biosynthetic Potency of Taxol from Fungal Endophytes of Conifers Plants; Genome Mining and Metabolic Manipulation.

Endophytic fungi have been considered as a repertoire for bioactive secondary metabolites with potential application in medicine, agriculture and food industry. The biosynthetic pathways by fungal endophytes raise the argument of acquisition of these machineries of such complex metabolites from the plant host. Diterpenoids "Taxol" is the most effective anticancer drug with highest annual sale, since its discovery in 1970 from the Pacific yew tree, Taxus brevifolia. However, the lower yield of Taxol from this natural source (bark of T. brevifolia), availability and vulnerability of this plant to unpredicted fluctuation with the ecological and environmental conditions are the challenges. Endophytic fungi from Taxus spp. opened a new avenue for industrial Taxol production due to their fast growth, cost effectiveness, independence on climatic changes, feasibility of genetic manipulation. However, the anticipation of endophytic fungi for industrial Taxol production has been challenged by the loss of its productivity, due to the metabolic reprograming of cells, downregulating the expression of its encoding genes with subculturing and storage. Thus, the objectives of this review were to (1) Nominate the endophytic fungal isolates with the Taxol producing potency from Taxaceae and Podocarpaceae; (2) Emphasize the different approaches such as molecular manipulation, cultural optimization, co-cultivation for enhancing the Taxol productivities; (3) Accentuate the genome mining of the rate-limiting enzymes for rapid screening the Taxol biosynthetic machinery; (4) Triggering the silenced rate-limiting genes and transcriptional factors to activates the biosynthetic gene cluster of Taxol.

Conjugation of Aspergillus flavipes Taxol with Porphyrin Increases the Anticancer Activity of Taxol and Ameliorates Its Cytotoxic Effects.

Taxol is one of the potential anticancer drugs; however, the yield of Taxol and its cytotoxicity are common challenges. Thus, manipulating the Taxol biosynthetic pathway from endophytic fungi, in addition to chemical modification with biocompatible polymers, is the challenge. Four fungal isolates, namely, Aspergillus flavipes, A. terreus, A. flavus, and A. parasiticus, were selected from our previous study as potential Taxol producers, and their potency for Taxol production was evaluated in response to fluconazole and silver nitrate. A higher Taxol yield was reported in the cultures of A. flavipes (185 µg/L) and A. terreus (66 µg/L). With addition of fluconazole, the yield of Taxol was increased 1.8 and 1.2-fold for A. flavipes and A. terreus, respectively, confirming the inhibition of sterol biosynthesis and redirecting the geranyl phosphate pool to terpenoids synthesis. A significant inhibition of ergosterol biosynthesis by A. flavipes with addition of fluconazole was observed, correlating with the increase on Taxol yield. To increase the Taxol solubility and to reduce its cytotoxicity, Taxol was modified via chemical conjugation with porphyrin, and the degree of conjugation was checked from the Thin layer chromatography and UV spectral analysis. The antiproliferative activity of native and modified Taxol conjugates was evaluated; upon porphyrin conjugation, the activity of Taxol towards HepG2 was increased 1.5-fold, while its cytotoxicity to VERO cells was reduced 3-fold.

Policosanol as a new inhibitor candidate for vascular calcification in diabetic hyperlipidemic rats.

This work mainly aimed to investigate the probable changes of aortic calcification by policosanol, omega-3 fatty acids in comparison with atorvastatin and subsequent progression of atherosclerosis in diabetic hyperlipemic rat model. Adult male albino rats of wistar strain (30) were divided into five groups (n = 6/group); one was fed normal diet and was used as a normal group, the other groups received alloxan, atherogenic diet (CCT - rat chow diet supplemented with 4% cholesterol, 1% cholic acid, and 0.5% thiouracil) and categorized as follows: the second group received no treatment and kept as control (diabetic hyperlipidemic control group (DHC)). The other groups received daily oral doses of atorvastatin, policosanol (10 mg/kg body weight) and ω-3 (50 mg/kg body weight), respectively, for eight weeks. Different biomarkers were used for the evaluation that included inflammatory (C reactive protein (CRP), tumor necrosis factor α (TNF-α)), oxidative stress (glutathione (GSH), malondialdehyde (MDA)) bone calcification markers (alkaline phosphatase (ALP), Vitamin D, parathyroid hormone (PTH)), lipogram pattern in addition to histochemical demonstration of calcium in the aorta. Diabetic hyperlipemic group demonstrated significant hyperglycemia, hyperlipidemia, and increased inflammation, oxidative stress, calcification, and finally atherogenesis progression. Treatment of diabetic hyperlipemic rats with, policosanol, omega-3 fatty acids (natural products) and atorvastatin for eight weeks significantly increased high-density lipoprotein cholesterol (HDL-C), Vitamin D, decreased aortic vacuoles number, and inhibited calcification process. Policosanol induced more remarkable reduction in the density and number of foam cells and improved the intimal lesions of the aorta as compared to atorvastatin. Drugs under study exerted hypoglycemic effect along with an inhibition of inflammation, oxidative stress, and calcium deposition with certain variations but policosanol effect was remarkable in comparison with other drugs.