Anti-leukemic principle(s) from Momordica charantia seeds induce differentiation of HL-60 cells through ERK/MAPK signalling pathway.

Myeloid leukemia is one of the major causes of deaths among elderly with very poor prognosis. Due to the adverse effects of existing chemotherapeutic agents, plant-derived components are being screened for their anti-leukemic potential. Momordica charantia (bitter gourd) possesses a variety of therapeutic activities. We have earlier demonstrated anti-leukemic activity of acetone extract of M. charantia seeds. The present study reports purification of differentiation inducing principle(s) from further fractionated seed extract (hexane fraction of the acetone extract, Mc2-Ac-hex) using HL-60 cells. Out of the 5 peak fractions (P1-P5) obtained from normal phase HPLC of the Mc2-Ac-hex, only peak fraction 3 (P3) induced differentiation of HL-60 cells as evident from an increase in NBT-positive cells and increased expression of cell surface marker CD11b. The P3 differentiated the HL-60 cells to granulocytic lineage, established by increased CD15 (granulocytic cell surface marker) expression in the treated cells. Further, possible molecular mechanism and the signalling pathway involved in the differentiation of HL-60 cells were also investigated. Use of specific signalling pathway inhibitors in the differentiation study, and proteome array analysis of the treated cells collectively revealed the involvement the of ERK/MAPK mediated pathway. Partial characterization of the P3 by GC-MS analysis revealed the presence of dibutyl phthalate, and derivatives of 2,5-dihydrofuran to be the highest among the 5 identified compounds. This study thus demonstrated that purified differentiation-inducing principle(s) from M. charantia seed extract induce HL-60 cells to granulocytic lineage through ERK/MAPK signalling pathway. Supplementary Information: The online version contains supplementary material available at 10.1007/s10616-022-00547-x.

Securinine Induces Differentiation of Human Promyelocytic Leukemic HL-60 Cells through JNK-Mediated Signaling Pathway.

Acute myeloid leukemia is characterized by abnormal differentiation of hematopoietic stem cells, leading to the accumulation of immature myeloid cells. Differentiation therapy has been a successful treatment option for acute promyelocytic leukemia but suffers from adverse effects. Therefore, search for novel differentiation-inducing agents with minimal side effects is desirable. Securinine, a naturally-occurring alkaloid, induces differentiation in various leukemic cells and apoptosis in other types of cancers. However, the underlying molecular mechanism(s) remain elusive. Our study aimed to elucidate the possible molecular mechanism(s) and signaling events involved in securinine-induced differentiation of HL-60 cells. Securinine inhibited proliferation in a time- and dose-dependent manner and triggered differentiation. A higher CD14+ population indicated maturation toward monocytic lineage. Securinine caused cell cycle arrest at the G0/G1 phase and enhanced ROS generation. Quantitative gene expression analysis showed significant down-regulation of C/EBP-α, C/EBP-ε, GAΤΑ, and c-myc and up-regulation of the PU.1 gene. The expression of distinct protein kinases Lyn, Chk-2, Yes, FAK, c-Jun, and JNK were enhanced. Use of specific inhibitors of crucial intracellular signaling proteins indicated that JNK and ERK blockade resulted in a significant decline in differentiation. These data thus confirm that securinine induces differentiation through the activation of the JNK-ERK signaling pathway in HL-60 cells.

Flavonoids modulate tight junction barrier functions in hyperglycemic human intestinal Caco-2 cells.

OBJECTIVES: Diabetes mellitus is a chronic disease requiring lifelong medical attention. With hundreds of millions suffering worldwide and a rapidly rising incidence, diabetes mellitus poses a great burden on health care systems. Recent studies investigating the underlying mechanisms involved in disease development in diabetes point to the role of the dysregulation of the intestinal barrier. Hyperglycemia-mediated tight junction deformity is known to contribute to leaky gut in various metabolic disorders. The present study aimed to investigate the role of oxidative stress on intestinal epithelial tight junction (TJ) barrier functions in hyperglycemia. Because many flavonoids are known to influence the cellular redox state, exploring these flavonoids may help to understand the role of TJ barrier in hyperglycemia-mediated oxidative stress, which in turn might unfold the association of oxidative stress and dysfunction of barrier-forming TJs. METHODS: Caco-2 cells were stimulated with high glucose (HG), with or without flavonoids (quercetin, morin, naringenin), for 24 h. We determined cellular viability, levels of reactive oxygen species, and mitochondrial membrane potential in flavonoids treated HG-Caco-2 cells. The levels of the proinflammatory cytokines, glucose uptake, and expression of glucose transporters were determined on flavonoids treatment. We investigated the effect of flavonoids on TJs functions by measuring transepithelial electrical resistance (a TJ integrity marker), membrane permeability using tracer compounds, and the expressions levels of TJs related molecules on hyperglycemic Caco-2 cell monolayers. RESULTS: We found that high glucose treatment resulted in reduced cell viability, increased reactive oxygen species production, measurable mitochondrial dysfunction, and decreased transepithelial electrical resistance, with increased membrane permeability. Treatment with the test flavonoids produced increased cell viability and reduced glucose uptake of HG-Caco-2 cells. A concomitant decrease in reactive oxygen species production, proinflammatory cytokines, and Glut-associated genes and proteins were identified with flavonoid treatment. Flavonoids prevented derangement of TJs protein interaction and stabilized membrane permeability. CONCLUSIONS: These findings indicate that flavonoids confer protection against hyperglycemia-mediated oxidative stress and enhance intestinal barrier functions by modulating underlying intracellular molecular mechanisms.