Anticancer effect of Piper betle leaf extract on KB cell lines – an in vitro study

Background: Various researchers have stated a causal association of betle quid chewing with oral cancer and other potentially malignant disorders of oral cavity. On the contrary, Piper betle leaf when used alone has potential medicinal benefits including anticancer, anti-helminthic, hepato-protective and antioxidant activities. In this is study we examined the anti-cancer activity of Piper betle extract (aqueous) on KB- cancer cell lines Aims: To observe the anti- cancer activity of Piper betle leaf extract on KB cancer cell lines. Setting and Design: The study was conducted in Biogenix Research Centre, Thiruvananthapuram. The KB cancer cell lines were procured from NCCS, Pune. Methods and Material: The cancer cell lines were treated with increasing concentration of Piper betle leaf extract 6.25,12,25,50 & 100?g/ml. The cytotoxic effect of the extract on the cells was studied by physical indicators of cytotoxic changes by observing the cells under an inverted phase contrast microscope, for any detectable changes in the cell morphology and by MTT assay method to assess the percentage of viability of cells. Results: The cancer cells showed considerable changes in the cell morphology suggestive of cell cytotoxicity and apoptosis after the treatment with the extract. The results of the MTT assay showed that the percentage viability of the cancer cells decreased with increasing concentrations of the extract, The percentage of viability of cells was noted to be 43.42% with the highest concentration of 100?g/ml of Piper betle leaf extract which proves that Piper betle leaf extract has anticancer activity. Conclusion: The cytotoxic potential of Piper betle leaf may be used to develop chemotherapeutic agent, but further focused studies of anticancer properties and isolation of compounds from Piper betle leaf are necessary to prove its worth in the cancer therapy.

Probiotics study with Streptococcus salivarius and its ability to produce bacteriocins and adherence to KB cells / Estudo de probióticos Streptococcus salivarius e sua habilidade de produzir bacteriocinas e aderência à células KB

Abstract Introduction Streptococcus salivarius is a dominant oral species and the best suitable candidate for probiotic of the oral cavity. Since Streptococcus salivarius is able to produce bacteriocins against Streptococcus pyogenes interest has been focused on the use of it as a probiotic to avoid sore throats by Streptococcus pyogenes. Objective This study is for selecting Streptococcus salivarius strains for potential use as probiotics for the oral mucosa, that is, production of bacteriocin against Streptococcus pyogenes and the ability to bind to KB cells. Material and method Tongue material from 45 students was collected and seeded on Mitis Salivarius Agar plaques. The strains were tested by the production of bacteriocin-like substances (BLIS) against S. pyogenes, biochemically and PCR for identification of S. salivarius. The best strains were tested for adherence to KB cells. Briefly, S. salivarius strains were cultured in broth, washed and suspended at 108cells/ml. KB cells were inoculated into plaques, washed and incubated with the bacteria, for adhesion. These were washed for lysis of the KB cells and release bacteria for determination of CFU. Result The bacteriocin test showed that 133 strains presented inhibition of S. pyogenes. The samples tested for adhesion to KB cells, presented different profiles and only three strains presenting high adhesion capacity. Conclusion The selection of strains of Streptococcus salivarius with high inhibitory activity against Streptococcus pyogenes, as well as adherence to KB cells leads us to the next future step, that is, to use the best strains for in vivo colonization tests

Resumo Introdução Streptococcus salivarius é uma espécie dominante na cavidade bucal e tem sido indicada como um ótimo candidato para uso como probiótico. Visto que a espécie Streptococcus salivarius é capaz de produzir bacteriocinas contra Streptococcus pyogenes, desenvolveu-se interesse no uso desse microrganismo como probiótico, para evitar amigdalites causadas por Streptococcus pyogenes. Objetivo A pesquisa em questão tem o objetivo de selecionar cepas de Streptococcus salivarius para seu uso potencial como probióticos na cavidade bucal, ou seja, produção de bacteriocinas contra Streptococcus pyogenes e habilidade de aderência à células KB. Material e método Coletou-se material de língua de 45 estudantes e semeou-se em placas de ágar Mitis Salivarius. As amostras foram testadas para verificar a produção de substâncias semelhantes à bacteriocina (BLIS) contra S. pyogenes, bioquimicamente e através de PCR para identificação de S. salivarius. As melhores cepas foram testadas quanto aderência à células KB. Resumidamente, as cepas de S. salivarius foram cultivadas em caldo, lavadas e suspensas à correspondência de 108 cels/ml. As células KB foram inoculadas em placas, lavadas e incubadas com as bactérias, para adesão. Estas foram lavadas para lise das células KB e liberação das bactérias para determinação de UFC. Resultado O teste de bacteriocina, mostrou que 133 cepas apresentaram atividade inibitória contra Streptococcus pyogenes. As cepas testadas para aderência à células KB, apresentaram diferentes perfis e somente três com alta capacidade de adesão. Conclusão: A seleção de cepas de Streptococcus salivarius com alta atividade inibitória contra Streptococcus pyogenes, bem como aderência a células KB, pode nos levar ao próximo passo, ou seja, o uso das melhores cepas para o estudo de colonização in vivo.

Cytotoxic and apoptotic potential of Myristica fragrans Houtt. (mace) extract on human oral epidermal carcinoma KB cell lines

Several studies have revealed that certain naturally occurring medicinal plants inhibit the growth of various cancers. The present study was conducted to evaluate cytotoxicity and apoptotic induction potential of Myristica fragrans Houtt mace extract. The cytotoxic activity of the Myristica fragrans Houtt mace acetone extract was assayed by MTT assay on human oral epidermal carcinoma KB cell lines. KB cells were incubated with different concentration of mace extract ranging from 25 to 125 µg/mL for 24hrs. The apoptotic induction potential was also studied by the analysis of Bcl-2 protein and gene expression in mace extract incubated KB cell lines using western blotting technique and real-time polymerase chain reaction. The mace extract exhibited cytotoxicity and anticancer effect against KB cell lines and it also suppressed the growth of cancer cells, therefore growth inhibitory effect was noted in extract treated cell lines. The apoptotic potential of mace extract was accompanied by reduced gene expression of Bcl-2 compared to the untreated KB cells. The mace extract shows the cytotoxic activity and induced the apoptosis through the modulation of its target genes Bcl-2 in the KB cell lines, suggesting the potential of mace as a candidate for oral cancer chemoprevention. This can be further investigated in vivo for its anticancer potential.

The effects of smoking-individual-serum on the invasion of F.Nucleatum to KB cells and MMP-1 expres-sion of KB cells / 实用口腔医学杂志

Objective:To investigate the effects of serum from smoking individuals with periodontitis in the process of Fusobocterium.Nucleatum(Fn)invading KB cells and the expression of matrixmetalloproteinase1(MMP1)of KB cells.Methods:Serum was prepared from 10 smokers with periodontitis(group Y),10 nonsmokers with periodontitis(group N)and 5 periodontal healthy nonsmokers(group H).Serum of 200,400 and 800 μl from the subjects was added to the model of Fn invading KB cells respectively and cultured for 24 hours,the cfu of cellinvaded bacteria was estimated by colony counting.MMP1 protein level in culture supernatant wasmeasured by ELISA.Results:In the 800 μl serum groups,the percentage of invaded cfu was 12.59 ±1.27,8.03 ±0.075 and 7.99±0.14 in group Y,N and H(P ＜0.05)respectively,the concentration(μg/L)of MMP1 in the cultrue supernatant of group Y,Nand H was 400.04 ±21.02,252.57 ±18.89 and 262.47 ±35.29(P ＜0.05)respectively.In the 200 μl and 400 μl serum groups,no significant difference was detected in invasion cfu of Fn in KB cells and MMP1 secrition of KB cells among Y,N and H groups(P>0.05).Conclusion:Higher doses of smokingserum might play a role in the course of Fn invading KB cells and promote the expression of MMP1 secretion from KB cells.

Reversal Effect of A Novel N-sugar Substituted Thalidomide Analogue on Multidrug Resistant Human Nasopharyngeal Carcinoma Cells / 生物化学与生物物理进展

One major problem to successful treatment of cancer is the development of resistance by tumor cells to multiple chemotherapeutic drugs, a phenomenon named multidrug resistance (MDR). Searching for the novel chemotherapeutical agents is one of the important strategies for overcoming MDR. By using a cytotoxicity assay, flow cytometry analysis, Western-blotting and RT-PCR, a drug (Taxol, TAX) resistant human nasopharyngeal carcinoma KB cell line (KB/TAX) was established by addition of the drug to the cell cultures gradually, then a novel N-sugar substituted thalidomide analogue (STA-35) was investigated for its reversal effect on MDR of KB/TAX cells and possible mechanism. The results showed that KB/TAX cells were resistant to several chemotherapeutical agents, and the relative resistance to TAX was 73.1. Compared with parental KB cells, the function and protein expression of P-glycoprotein (P-gp), as well as mdr1 gene in the KB/TAX cells were remarkable reduced. Moreover, both KB and KB/TAX cells were sensitive to STA-35, the relative resistance to TAX on KB/TAX cells was decreased by the addition of STA-35. Furthermore, STA-35 (5 ～20 ?mol/L)was capable to reduced the activity of P-gp by increasing the accumulation of rhodamine 123, decreasing P-gp expression in KB/TAX cells in a dose dependent manner , but had no effect on the mdr1 gene expression. These results suggest a potential action of STA-35 as MDR reversing agent, and one of the possible mechanisms could be the suppression of P-gp function and protein expression.

Adhesion of Weissella cibaria to the Epithelial Cells and Factors Affecting its Adhesion

We evaluated the ability of lactic acid bacteria, Weissella cibaria, isolated from the oral cavity to adhere to epithelial cells. W. cibaria efficiently adhered to KB cells and HeLa cells. In addition, W. cibaria efficiently adhered to Fusobacterium nucleatum. But the adhesiveness of W. cibaria disappeared upon exposure to LiCl or pronase, suggesting that the S-layer proteins of W. cibaria mediated the adhesiveness. The molecular mass of the S-layer proteins extracted from W. cibaria was approximately 50 kDa. When W. cibaria strains were washed with 0.45% saline, the bacteria were efficiently adhered to the epithelial cells. In conclusion, W. cibaria has the ability to adhere to epithelial cells through the S-layer proteins.

A study of the Ca2+ and the apoptosis of the KB cell lines after 10 Gy irradiation / 대한구강악안면방사선학회지

PURPOSE: Ionizing radiations have been reported as an apoptosis initiating stimulus in various cells and it has established that sustained elevations in [Ca2+] can lead to DNA fragmentation by Ca2+-dependent endonucleases, ultimately resulting in apoptotic cell death. The previous experiments have been reported by using primarily thymocytes and lymphocytes and the change of [Ca2+] was measured only by minutes or hours respectively. We need to evaluate [Ca2+] in both several minutes and hours after irradiation of radiation of radiation therapy and verify the apoptotic cells. MATERIALS AND METHODS: We have measured [Ca2+] in human gingival epitheloid cancer cell with 10 Gy irradiation, at minutely intervals and hourly intervals using digitized video-intensified fluorescence microscopy and the fluorescent Ca2+ indicator dye, fura-2. In order to find out that the transient rise in [Ca2+] could induced apoptosis, cells were incubated for 1 hour at 37 degrees C with TdT enzyme, rinsed and resuspended containing fluorescence and observed under a confocal fluorescence microscope. MTT assay was done to determine cell activity and LDH assay was done to determine the amount of necrotic cells. RESULTS: After irradiation, the transient and temporal increasing of [Ca2+] in the KB cells was founded. Though, there was no change in the intracellular [Ca2+] at 30 minutes and 2 hours after irradiation. We could detect of DNA fragmented cells at 4 hours after 10 Gy irradiated cells. There were no significant differences between 4 hour, 1 day, 3 day cells. There were no significant differences in MTT and LDH assay between the irradiated group and the control group after 4 hours and 1 day. Though after 3 days there were differences in MTT and LDH assay between the irradiated group was significantly decreased than the control group, in LDH assay the number of necrotic cell death of the irradiated was higher than the control group. CONCLUSION: In KB cells there were incipient and temporal increasing of the [Ca2+] with 10 Gy irradiation and the apoptosis was founded from 4 hours later which was earlier than seeing of the change of the amount of the cellular ability and necrosis.