Background: β-caryophyllene (BCP) is a naturally occurring bicyclic sesquiterpene extracted from various plants, and widely used as a medicinal agent for various diseases. During hepatocellular carcinoma (HCC) development, cancer cells generally exhibit increased cell proliferation due to mutations or aberrant expression of key regulatory genes. The current study determines the cytotoxic effects of BCP alone or in combination with doxorubicin (DOX) and cisplatin (DDP) on HCC cells, and elucidates the underlying mechanism of BCP to exert its anticancer activities.Materials and methods: HepG2, SMMC-7721 HCC cells, and HL-7702 normal liver cells were treated with BCP, DOX, and DDP individually or combinatorially. Cell proliferation assay, flow cytometric assay, and Western blot were employed to evaluate the cytotoxic effects of these treatments. Transwell assays were used to examine BCP’s effects on HCC cell migration and invasion. RNA-seq analysis was used to determine BCP’s primary target genes in HepG2 cells. Integrative analysis of differentially expressed genes (DEGs) of RNA-seq data with an HCC TCGA dataset identified BCP-targeted genes that were verified by RT-qPCR analysis. Ectopic gene expression, cell viability, and colony formation assay were performed to validate the primary targets of BCP.Results: BCP selectively inhibited HCC cell proliferation while exhibited relatively low toxicity in normal liver cells; however, DOX and DDP showed higher toxicity in normal cells than that in HCC cells. In combinatorial treatments, BCP synergistically enhanced cytotoxicity of DOX and DDP in HCC cells but this effect was markedly reduced in HL-7702 cells. BCP treatment reduced migration and invasion of HCC cells. Furthermore, RNA-seq analyses of BCP-treated HepG2 cells identified 433 protein-coding DEGs. Integrative analyses revealed five BCP-targeted DEGs regulating the MAPK signaling pathway. Among these five genes, three displayed a significantly positive correlation of their expression with the overall survival of HCC patients. As a primary target, PGF was significantly downregulated by BCP treatment, and its exogenous expression desensitized HCC cells to BCP-mediated inhibition.Discussion: BCP inhibits malignant properties of HCC and synergistically sensitizes the anticancer activity of DOX and DDP. In HCC cells, BCP primarily targets the PGF gene and MAPK signaling pathway.

The increased pressures generated by human activities in form of climate change and habitat fragmentation have caused serious threats for Sindh ibex (Capra aegagrus blythii). Balochistan, recognized as one of the most vulnerable regions to climate change in Pakistan, has limited data on the distribution and population status of this species. To address this gap, our study aims to assess the spatial and temporal distribution and population trend of Sindh ibex in the Khuzdar and Lasbela districts of Balochistan. The study covered an area of approximately 45,123.97 km². It involved a double-observer survey at ten sites from 2019 to 2022, using the BBRecapture package in the R programming language to estimate ibex population trend and dispersion. We provided a comprehensive update of the species geographic range, along with estimates of its current abundance and population trends. Over the course of four years, the ibex population fluctuated. The ibexes detected dropped from 720 (male to female ratio 1:1.21) and 710 (male to female ratio 1:1.08) in 2019 and 2020, respectively, to 550 (male to female ratio 1:1.35) and 548 (male to female ratio 1:1.09). Overall, this study provides valuable insights into the population trends, gender ratios, habitat preferences, and density of Sindh ibex in Balochistan over the four-year period. These findings contribute significantly to the baseline data on the species ecology and will further help in the conservation of these species at local scale.

Livestock herding is a vital practice in Balochistan, contributing to the economy and culture. The livestock sector is significant in Balochistan, providing 20% of the national stock. Large predators and their prey species, including livestock, have coexisted in these mountainous landscapes for centuries. The aim of the present research is to investigate the impacts of livestock depredation by large predators on livelihoods and predator conservation in two districts of Balochistan, Pakistan. A human–carnivore conflict survey was conducted from July to September 2019, collecting data from 311 residents in a selected study area. Large predators in the study area preyed on a total of 876 livestock during a one-year period, including 560 goats, 292 sheep, 19 cows, and 5 donkeys. The gray wolf is the leading predator, responsible for 66.3% of livestock depredation, followed by the caracal (24.3%), Asiatic jackal (8.9%), and striped hyena (0.6%). The total economic loss was USD 78,694. Overall, 80% of respondents had a negative perception of wolves compared to 24.4% for caracals. Only 20.6% of respondents knew about the importance of conserving carnivores. Livestock depredation by carnivores in the study area created a negative perception of these animals among people. There is a lack of awareness about the importance of conserving carnivore species and their role in the ecosystem. This lack of understanding has ultimately led to detrimental effects on predator populations. It is imperative to raise awareness among people about the ecological significance of carnivores through community meetings, seminars in educational institutions, and providing basic education to herders about effective livestock guarding practices.