Validation of a Quantification Method for Curcumin Derivatives and Their Hepatoprotective Effects on Nonalcoholic Fatty Liver Disease

Curcumin (CM), demethoxycurcumin (DMC), and bisdemethoxycurcumin (BDMC) are major curcumin derivatives found in the rhizome of turmeric (Curcuma longa L.), and have yielded impressive properties to halt various diseases. In the present study, we carried out a method validation for curcumin derivatives and analyzed the contents simultaneously using HPLC with UV detection. For validation, HPLC was used to estimate linearity, range, specificity, accuracy, precision, limit of detection (LOD), and limit of quantification (LOQ). Results showed a high linearity of the calibration curve, with a coefficient of correlation (R2) for CM, DMC, and BDMC of 0.9999, 0.9999, and 0.9997, respectively. The LOD values for CM, DMC, and BDMC were 1.16, 1.03, and 2.53 ng/μL and LOQ values were 3.50, 3.11, and 7.67 ng/μL, respectively. Moreover, to evaluate the ability of curcumin derivatives to reduce liver lipogenesis and compare curcumin derivatives’ therapeutic effects, a HepG2 cell model was established to analyze their hepatoprotective properties. Regarding the in vivo study, we investigated the effect of DMC, CM, and BDMC on nonalcoholic fatty liver disease (NAFLD) caused by a methionine choline deficient (MCD)-diet in the C57BL/6J mice model. From the in vitro and in vivo results, curcumin derivatives alleviated MCD-diet-induced lipid accumulation as well as high triglyceride (TG) and total cholesterol (TC) levels, and the protein and gene expression of the transcription factors related to liver adipogenesis were suppressed. Furthermore, in MCD-diet mice, curcumin derivatives suppressed the upregulation of toll-like receptors (TLRs) and the production of pro-inflammatory cytokines. In conclusion, our findings indicated that all of the three curcuminoids exerted a hepatoprotective effect in the HepG2 cell model and the MCD-diet-induced NAFLD model, suggesting a potential for curcuminoids derived from turmeric as novel therapeutic agents for NAFLD.

Genetic Profile Evaluation of Human Cell Lines Treated with Anastatica hierochuntica Using Forensic DNA Fingerprinting Markers

Cell line authentication using Short Tandem Repeats (STRs) is necessary to ensure the integrity of the cell for its continuous culture and to identify misidentification and cross-contamination issues. This study investigates the changes in the genetic profile of MCF-7 and HepG2 cell lines caused by the methanolic leaf extract of Anastatica hierochuntica (AH) using human identification based STR markers. MCF-7 and HepG2 cell lines were treated with various concentrations of AH extracts for three different periods. The treated and control cells' DNA was extracted using a QIAamp® DNA Micro Kit, quantified using a Quantifiler Duo DNA Quantification Kit, and amplified using an AmpFlSTR Identifiler plus PCR Amplification Kit. The concentrations of the DNA extracted from control and MCF-7 and HepG2 cell lines treated with AH extract at 300 to 2400 µg/ml for 24hr and 150 to 2400 µg/ml for 48 and 72hrs were statistically significant (p<0.05). Microsatellite instability (MSI), loss of heterozygosity (LOH), insertion/deletions changes in the STRs profile were observed in treated cell lines at 1200 and 2400 µg/ml in MCF-7 cells for 48 and 72hrs and HepG2 cells for 24, 48, and 72hrs. We conclude that the highest concentration of AH extracts affected the genotype of the cell lines leading to misidentification. Therefore, cell line authentication by forensic DNA analysis techniques plays a decisive role for cells tested with a high concentration of chemical compounds and gives the forensic investigator an insight into these changes in the STR genotype of a victim/suspect who has been been under long term chemotherapeutic treatment.

Evaluation of Antioxidant and Antidiabetic Potential of Polyherbal Drug Made with Three Herbs: An In vitro Cell Culture Model

In diabetes, the postprandial phase is characterized by a rapid and large increase in blood glucose levels, and the possibility that the postprandial “hyperglycemic spikes” may be relevant to the onset of cardiovascular complications has recently received much attention. Medicinal use of herbal medicine in the treatment and prevention of diseases including diabetes has a long history compared to conventional medicine. These plants have no side effects and many existing medicines are derived from the plants. Hence, the current investigation was planned to make a poly herbal drug (PHD) through Punica granatum (fruits), Illicium verum (flowers) and Nyctanthes arbor (leaves) and assess their antioxidant and antidiabetic activities in vitro and in HepG2 cell line. The respective plant methanolic extracts and PHD are exposed to phytochemical assessment and to discriminate the bioactive factors by Gas Chromatography–Mass Spectrometry. We evaluated the antioxidant properties 2, 2-diphenyl-1-picrylhydrazyl scavenging, hydrogen peroxide scavenging, thiobarbituric acid reactive substances and total antioxidant activity of individual plant extracts and the PHD. At the same time, In vitro and cell culture approaches were used to assess the anti-diabetic activity. The PHD has a higher concentration of secondary metabolites than individual plant extracts, according to our findings. On the other hand, we also notice that PHD demonstrated higher antioxidant capability and considerable in vitro glucose-lowering effects along with noteworthy inhibition of ɑ-amylase, glucosidase, lipase, dipeptidyl peptidase-IV, collagenase and protein glycation in HepG2 cell line. In conclusion, this study clearly demonstrated the significant antioxidant and antidiabetic activities of the PHD. Hence, PHD may be used as a potential source in the management of diabetes, hyperglycemia and the related state of oxidative stress.

The Curious Case of the HepG2 Cell Line: 40 Years of Expertise

Liver cancer is the third leading cause of cancer death worldwide. Representing such a dramatic impact on our lives, liver cancer is a significant public health concern. Sustainable and reliable methods for preventing and treating liver cancer require fundamental research on its molecular mechanisms. Cell lines are treated as in vitro equivalents of tumor tissues, making them a must-have for basic research on the nature of cancer. According to recent discoveries, certified cell lines retain most genetic properties of the original tumor and mimic its microenvironment. On the other hand, modern technologies allowing the deepest level of detail in omics landscapes have shown significant differences even between samples of the same cell line due to cross- and mycoplasma infection. This and other observations suggest that, in some cases, cell cultures are not suitable as cancer models, with limited predictive value for the effectiveness of new treatments. HepG2 is a popular hepatic cell line. It is used in a wide range of studies, from the oncogenesis to the cytotoxicity of substances on the liver. In this regard, we set out to collect up-to-date information on the HepG2 cell line to assess whether the level of heterogeneity of the cell line allows in vitro biomedical studies as a model with guaranteed production and quality.