Luolin Wang,1,* Zhenglei Xu,1,* Aarti Bains,2 Nemat Ali,3 Zifang Shang,4 Abhinandan Patil,5 Sandip Patil6 

1Department of Gastroenterology, Shenzhen People’s Hospital (The Second Clinical Medical College, Jinan University; The First Affiliated Hospital, Southern University of Science and Technology), Shenzhen, Guangdong, People’s Republic of China; 2Deparment of Microbiology, Lovely Professional University, Phagwara, Punjab, India; 3Department of Pharmacology and Toxicology, College of Pharmacy, King Saud University, Riyadh, Saudi Arabia; 4Research Experiment Center, Meizhou People’s Hospital, Meizhou Academy of Medical Sciences, Meizhou, Guangdong, People’s Republic of China; 5Department of Pharmaceutics, D. Y. Patil Education Society (Deemed to be University), Kolhapur, Maharashtra, India; 6Department of Haematology and Oncology, Shenzhen Children’s Hospital, Shenzhen, Guangdong, People’s Republic of China

*These authors contributed equally to this work

Correspondence: Abhinandan Patil, Department of Pharmaceutics, D. Y. Patil Education Society (Deemed to be University), Kadamwadi, Kolhapur, Maharashtra, India, Email abhisirdyp@gmail.com Sandip Patil, Department of Haematology and Oncology, Shenzhen Children’s Hospital, Shenzhen, Guangdong, People’s Republic of China, Email sandippatil1309@yahoo.com

Introduction: This study aims to systematically assess the anticancer potential of distinct Lactobacillus strains on Human Colorectal Tumor (HCT) 115 cancer cells, with a primary focus on the apoptotic mechanisms involved. Lactobacillus strains were isolated from sheep milk and underwent a meticulous microbial isolation process. Previous research indicates that certain probiotic bacteria, including Lactobacillus species, may exhibit anticancer properties through mechanisms such as apoptosis induction. However, there is limited understanding of how different Lactobacillus strains exert these effects on cancer cells and the underlying molecular pathways involved.
Methods: Cytotoxicity was evaluated through 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays and exposure durations of Lactobacillus cell-free lyophilized filtrates. Additional apoptotic features were characterized using 4.6-diamidino-2-phenylindole (DAPI) analysis for nuclear fragmentation and Annexin V/PI analysis for apoptosis quantification. Genetic analysis explored the modulation of apoptotic proteins (Bax and Bcl2) in response to Lactobacillus treatment. Whole-genome sequencing (WGS) was performed to understand the genetic makeup of the Lactobacillus strains used in the study.
Results: The study demonstrated a significant reduction in HCT 115 cell viability, particularly with L. plantarum, as evidenced by Sulforhodamine B (SRB) and MTT assays. DAPI analysis revealed nuclear fragmentation, emphasizing an apoptotic cell death mechanism. Annexin V/PI analysis supported this, showing a higher percentage of early and late apoptosis in L. plantarum-treated cells. Genetic analysis uncovered up-regulation of pro-apoptotic protein Bax and down-regulation of anti-apoptotic protein Bcl2 in response to Lactobacillus treatment. WGS study revealed a strain reported to NCBI PRJNA439183.
Discussion: L. plantarum emerged as a potent antiproliferative agent against HCT 115 cancer cells, inducing apoptosis through intricate molecular mechanisms. This study underscores the scientific basis for L. plantarum’s potential role in cancer therapeutics, highlighting its impact on antiproliferation, adhesion, and gene-protein regulation. Further research is warranted to elucidate the specific molecular pathways involved and to evaluate the therapeutic potential of L. plantarum in preclinical and clinical settings.

Keywords: sheep milk, Lactobacillus plantarum, cytotoxicity, apoptosis, human colorectal tumor 115, cancer therapeutics