Luteolin inhibits proliferation, triggers apoptosis and modulates Akt/mTOR and MAP kinase pathways in HeLa cells

Flavonoids, a subclass of polyphenols, have been shown to be effective against several types of cancer, by decreasing proliferation and inducing apoptosis. Therefore, the aim of the present study was to assess the anti-carcinogenic potential of luteolin on HeLa human cervical cancer cells, through the use of a cell viability assay, DNA fragmentation assay, mitochondrial membrane potential assay, cell cycle analysis using Annexin/PI staining and flow cytometry, gene expression analysis and a protein profiling array. Luteolin treatment exhibited cytotoxicity towards HeLa cells in a dose- and time-dependent manner, and its anti-proliferative properties were confirmed by accumulation of luteolin-treated cells in sub-G1 phases. Cytotoxicity induced by luteolin treatment resulted in apoptosis, which was mediated through depolarization of the mitochondrial membrane potential and DNA fragmentation. Furthermore, luteolin treatment increased the expression of various proapoptotic genes, including APAF1, BAX, BAD, BID, BOK, BAK1, TRADD, FADD, FAS, and Caspases 3 and 9, whereas the expression of anti-apoptotic genes, including NAIP, MCL-1 and BCL-2, was decreased. Cell cycle regulatory genes, including CCND1, 2 and 3, CCNE2, CDKN1A, CDKN2B, CDK4 and CDK2, were decreased following treatment. Expression of TRAILR2/DR5, TRAILR1/DR4, Fas/TNFRSF6/CD95 and TNFR1/TNFRSF1A, as well as pro-apoptotic proteins, including BAD, BAX and Cytochrome C were consistently increased, and the expression of antiapoptotic proteins, HIF1α, BCL-X, MCL1 and BCL2, were found to be decreased following treatment. Expression of AKT1 and 2, ELK1, PIK3C2A, PIK3C2B, MAPK14, MAP3K5, MAPK3 and MAPK1 was significantly decreased at the transcriptional level. Expression of GSK3b (p-ser9), PRAS 40 (p-Ther246), BAD (p-ser112), PTEN (p-ser380), AKT (p-ser473), ERK2 (p-Y185/Y187), RISK2 (p-ser386), P70S6k (p-Thr421/ser424), PDK1(p-ser241), ERK1 (p-T202/Y204) and MTOR (p-ser2448) was downregulated and expression of P53 (p-ser241) and P27(p-Thr198) was upregulated by luteolin in a dose-dependent manner, indicating its anti-proliferative and apoptosis enabling properties, and this may have been mediated via inhibition of the AKT and the MAPK pathways. © 2021 Spandidos Publications. All rights reserved

Luteolin Causes 5′CpG Demethylation of the Promoters of TSGs and Modulates the Aberrant Histone Modifications, Restoring the Expression of TSGs in Human Cancer Cells

Cancer progression is linked to abnormal epigenetic alterations such as DNA methylation and histone modifications. Since epigenetic alterations, unlike genetic changes, are heritable and reversible, they have been considered as interesting targets for cancer prevention and therapy by dietary compounds such as luteolin. In this study, epigenetic modulatory behaviour of luteolin was analysed on HeLa cells. Various assays including colony forming and migration assays, followed by biochemical assays of epigenetic enzymes including DNA methyltransferase, histone methyl transferase, histone acetyl transferase, and histone deacetylases assays were performed. Furthermore, global DNA methylation and methylation‐specific PCR for examining the methylation status of CpG promoters of various tumour suppressor genes (TSGs) and the expression of these TSGs at transcript and protein level were performed. It was observed that luteolin inhibited migration and colony formation in HeLa cells. It also modulated DNA methylation at promoters of TSGs and the enzymatic activity of DNMT, HDAC, HMT, and HAT and reduced the global DNA methylation. Decrease in methylation resulted in the reactivation of silenced tumour suppressor genes including FHIT, DAPK1, PTEN, CDH1, SOCS1, TIMPS, VHL, TP53, TP73, etc. Hence, luteolin‐targeted epigenetic alterations provide a promising approach for cancer prevention and intervention

Kaempferol Regresses Carcinogenesis through a Molecular Cross Talk Involved in Proliferation, Apoptosis and Inflammation on Human Cervical Cancer Cells, HeLa

Kaempferol, a flavonoid, contains a plethora of therapeutic properties and has demonstrated its efficacy against cancer. This study aims to unravel the molecular targets that are being modulated by kaempferol on HeLa cells. Various assays were performed, namely: MTT assay, flow cytometry to analyze DNA content and quantitate apoptosis. Quantitative PCR and protein profiling were performed to evaluate the modulated manifestation of different genes involved in apoptosis, cell growth and inflammation. Kaempferol exhibited reduction in cell viability of HeLa cells (IC50 = 50 µM 48 h), whereas it did not show any significant effect on viability of the AC-16 cell line. Kaempferol-impacted apoptosis was definitive, as it induced DNA fragmentation, caused disruption of membrane potential, accumulation of cells in the G2-M phase and augmented early apoptosis. Consistently, kaempferol induced apoptosis in HeLa cells by modulating the expression of various genes at both transcript and protein levels. It upregulated the expression of pro-apoptotic genes, including APAF1, BAX, BAD, Caspases 3, and 9, etc., at the transcript level and Bad, Bax, p27, p53, p21, Caspases 3 and 8 etc. at the protein level, while it downregulated the expression of pro-survival gene BCL-2, BIRC8, MCL-1, XIAP, and NAIP at the transcript level and Bcl-2, XIAP, Livin, clap-2 at the protein level. Kaempferol attenuated oxidative stress by upregulating GSH activity and anti-inflammatory response by suppressing NF-kB pathways. Moreover, kaempferol averted rampant cell division and induced apoptosis by modulating AKT/MTOR and MAP kinase pathways. Hence, kaempferol can be considered as a natural therapeutic agent with a differential profile

Phacoemulsification after penetrating keratoplasty with autologous limbal transplant and amniotic membrane transplant in chemical burns

We report a patient who had earlier penetrating keratoplasty with amniotic membrane transplant and autologous limbal cell transplant for chemical injury who underwent cataract surgery by phacoaspiration. A posterior limbal incision with corneal valve was made superotemporally with extreme caution to avoid damage to the limbal graft. Aspiration flow rates and vacuum were kept low to avoid any turbulence during surgery. A 6.0 mm optic diameter acrylic foldable intraocular lens was inserted in the bag. The patient achieved a best-corrected visual acuity of 6/12 at 10 months′ follow-up with a clear corneal graft. We conclude that caution during wound construction and phacoaspiration can help preserve corneal and limbal graft integrity in patients undergoing cataract surgery after corneal graft and limbal transplantation

Dasatinib in chronic myeloid leukemia: a limited Indian experience

Aim: To report our experience of the use of dasatinib in various phases of chronic myeloid leukemia (CML). Methods: Ten patients in various phases of CML, not responding to imatinib and started on dasatinib, were included and analyzed. The baseline characteristics of the patients and their salient features including the duration and response to initial therapy as well as to dasatinib, were noted. Results: Before starting dasatinib three patients were in chronic phase of CML while seven others were in the progressive phase (accelerated and blast phase) of CML. Half the patients developed transient grade 3 and 4 toxicities to dasatinib. Overall, the tolerability of the drug in all 10 patients was acceptable and none discontinued treatment. Three patients died due to progressive disease while the remaining seven are continuing the drug with the disease still under cytogenetic or hematological remission. Of the 10 patients, seven achieved complete hematological response and two of the accelerated phase/blast crisis patients achieved complete cytogenetic response. Overall, dasatinib was able to control disease for a median of 20.6 months. Conclusion: Despite small sample size and insufficient information on mutational analysis, dasatinib is effective in CML in INdia. Cost limits the use of second-generation tyrosine kinase inhibitors in India. Our observation is not suitable for survival analysis but the difference made by dasatinib in progressive disease and its tolerability needs to be acknowledged

Assessment of 285 cases of chronic lymphocytic leukemia seen at single large tertiary center in Northern India

Chronic lymphocytic leukemia (CLL) is the most common lymphoproliferative disorder in the West accounting for about 30% of leukemias, but about 2–4% in India. There is no large series reported from India, and hence we decided to undertake this study. We assessed the clinicohematological profiles and treatment outcomes in 285 patients seen over 11 years at our center (median age 59 years, 209 males). Sixty-three patients (22%) were asymptomatic and diagnosed incidentally. The median total leukocyte count at presentation was 50 × 10⁹/L. Rai stage distribution was: stage 0, 10%; stage I, 16%; stage II, 33%; stage III, 20%; and stage IV, 21%. Fifty percent of patients required treatment at presentation. Ninety-six patients received chlorambucil-based (overall response rate [ORR] 69%, complete remission [CR] 3%) and 27 patients received fludarabine-based (ORR 89%, CR 44%) chemotherapy. With a median follow-up of 2.9 years, the median overall survival (OS) and event-free survival (EFS) were 5.1 and 4.6 years, respectively. Fludarabine was more toxic, as half of the patients developed febrile neutropenia and various infections. The majority of patients presented with advanced stage disease. Fludarabine was less commonly used as first-line therapy. Advanced clinical stage (Rai III and IV) was associated with poor OS (hazard ratio [HR] 2.46, 95% confidence interval [CI] 1.19–5.11, p = 0.001) and EFS

Long-term outcomes for patients with acute myeloid leukemia: a single-center experience from AIIMS, India

Aim: To analyze clinicopathological characteristics of acute myeloid leukemia (AML) patients and to evaluate long-term outcome of these patients presented to single tertiary care center in India. Methods: We evaluated outcomes of 480 patients (age 8–60 years), classified into good, intermediate and poor risk according to cytogenetic results. Standard “3+7” induction therapy with dose of daunorubicin ranging from 45 to 90 mg/m2 followed by two to three courses of high-dose cytarabine (12–18 g/m2) as consolidation therapy was given to majority. Results: The complete remission rate of the treated population (407 patients) was 70% with 84.8% in good risk, 67.9% in intermediate risk and 54.2% in poor risk (P = 0.0001). Induction mortality was 18.4%. One hundred twenty-nine patients relapsed with median treatment free interval of 10.4 months. At a median follow-up of 34.5 months, the median overall survival (OS) was 20.6 months with an estimated 5-year survival rate of 35.5%. No difference was found in OS between the three risk groups; however, patients with intermediate risk had a better leukemia-free survival (LFS) in comparison to good risk. Multivariate analysis showed age, performance status, treatment completion and hematopoietic stem cell transplant affecting OS, while only treatment completion affected LFS. Conclusion: This is one of the largest single-center studies reflecting more accurately the outcome of AML in India. These results are likely due to uniform treatment protocols, intensification of induction and post-remission treatments with comprehensive supportive care