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      Evaluation of lymphocyte apoptosis in patients with oral cancer

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          Abstract

          Objectives

          To evaluate apoptotic levels of peripheral blood mononuclear cells (PBMCs) and apoptotic regulatory proteins (Bax and Bcl-2) in lymphocyte subsets of oral cancer (OC) patients and healthy controls (HC).

          Methodology

          The percentage of apoptotic cells and lymphocyte counts were measured in the first cohort using PBMCs obtained from 23 OC patients and 6 HC. In the second cohort, (OC, 33; HC, 13), the mean fluorescence intensity (MFI) of Bax and Bcl-2 in CD19 + B, CD4 + T, CD8 + T, and CD16 +56 + natural killer (NK) cells was determined via flow cytometry.

          Results

          The percentage of apoptotic cells was higher in the PBMCs of OC patients than in HC patients, particularly in patients with stage IV cancer ( p<0.05). However, lymphocyte counts were significantly lower in stage IV patients ( p<0.05). NK CD19 + B and CD16 +56 + cell counts were significantly lower in OC patients compared with HC patients ( p<0.001 and p<0.01, respectively), but CD4 + T cells were interestingly significantly higher in OC patients ( p<0.001). While Bax MFI was slightly higher, Bcl-2 MFI was significantly lower for all four lymphocyte subsets in OC samples, particularly in stage IV patients, when compared with HC. Consequently, Bax/Bcl-2 ratios showed an upward trend from HC to OC patients, particularly those in stage IV. We found similar trends in Bax and Bcl-2 MFI for tumor stage, tumor size, and lymph node involvement.

          Conclusions

          The increased lymphocyte apoptosis in stage IV OC patients may be related to higher Bax levels and lower Bcl-2 levels. The Bax/Bcl-2 ratio in lymphocytes may be useful to determine the prognosis of OC patients, and could be considered a mean for supportive treatment in the future.

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          Most cited references31

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          Lymphopenia as a prognostic factor for overall survival in advanced carcinomas, sarcomas, and lymphomas.

          Lymphopenia is frequent in advanced cancers and predicts the toxicity of chemotherapy. Its effect on relapse and survival is uncertain. Its prognostic value for survival was analyzed in three databases of previously reported prospective multicenter studies: (a) FEC chemotherapy in metastatic breast carcinoma; (b) CYVADIC in advanced soft tissue sarcoma (European Organization for Research and Treatment of Cancer-Soft Tissue and Bone Sarcoma Group 62791); and (c) prospective, consecutive phase III studies of aggressive diffuse large-cell non-Hodgkin's lymphomas conducted at Centre Léon Bérard between 1987 and 1993. Univariate and multivariate analyses of prognostic factors for survival were performed. The incidence of lymphopenia of 1, non-Hodgkin's lymphoma patients with international prognostic index (IPI) of > 0, and advanced soft tissue sarcoma and metastatic breast cancer patients with bone metastases. Inunivariate analysis, lymphopenia of <1,000/microL significantly correlated to overall survival in patients with metastatic breast cancer (median, 10 versus 14 mo; P < 0.0001), advanced soft tissue sarcoma (median, 5 versus 10 months; P < 0.01), and non-Hodgkin lymphoma (median, 11 versus 94 months; P < 0.0001). In multivariate analysis (Cox model), lymphopenia was an independent prognostic factor for overall survival in metastatic breast cancer [RR (relative risk), 1.8; 95% CI (confidence interval), 1.3-2.4] along with liver metastases and PS; in advanced soft tissue sarcoma (RR, 1.46; 95% CI, 1.0-2.1) along with liver metastases, lung metastases, and PS; and in non-Hodgkin's lymphoma (RR, 1.48; 95% CI, 1.03-2.1) along with IPI. Our findings show that lymphopenia is an independent prognostic factor for overall and progression-free survival in several cancers.
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            Mechanisms of action of Bcl-2 family proteins.

            The Bcl-2 family of proteins controls a critical step in commitment to apoptosis by regulating permeabilization of the mitochondrial outer membrane (MOM). The family is divided into three classes: multiregion proapoptotic proteins that directly permeabilize the MOM; BH3 proteins that directly or indirectly activate the pore-forming class members; and the antiapoptotic proteins that inhibit this process at several steps. Different experimental approaches have led to several models, each proposed to explain the interactions between Bcl-2 family proteins. The discovery that many of these interactions occur at or in membranes as well as in the cytoplasm, and are governed by the concentrations and relative binding affinities of the proteins, provides a new basis for rationalizing these models. Furthermore, these dynamic interactions cause conformational changes in the Bcl-2 proteins that modulate their apoptotic function, providing additional potential modes of regulation.
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              Tumor suppressor p53 is a regulator of bcl-2 and bax gene expression in vitro and in vivo.

              The p53 tumor suppressor gene product can induce apoptotic cell death through an unknown mechanism. Here we demonstrate that a temperature-sensitive p53 induces temperature-dependent decreases in the expression of the apoptosis-suppressing gene bcl-2 in the murine leukemia cell M1, while simultaneously stimulating increases in the expression of bax, a gene which encodes a dominant-inhibitor of the Bcl-2 protein. Mice deficient in p53 exhibit increases in Bcl-2 and decreases in Bax protein levels in several tissues as determined by immunohistochemical and immunoblot methods. The findings suggest a potential mechanism by which p53 regulates apoptosis, as well as responses to radiation and chemotherapeutic drugs in cancer.
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                Author and article information

                Journal
                J Appl Oral Sci
                J Appl Oral Sci
                jaos
                Journal of Applied Oral Science
                Faculdade De Odontologia De Bauru - USP
                1678-7757
                1678-7765
                07 September 2020
                2020
                : 28
                : e20200124
                Affiliations
                [1 ] orgnameChulalongkorn University orgdiv1Faculty of Medicine orgdiv2Department of Anatomy Bangkok Thailand originalChulalongkorn University, Faculty of Medicine, Department of Anatomy, Center of Excellence in Molecular Genetics of Cancer and Human Diseases, Bangkok, Thailand.
                [2 ] orgnameChulalongkorn University orgdiv1Faculty of Medicine orgdiv2Program of Medical Science Bangkok Thailand originalChulalongkorn University, Faculty of Medicine, Program of Medical Science, Bangkok, Thailand.
                [3 ] orgnameChulalongkorn University orgdiv1Faculty of Medicine orgdiv2Department of Medicine Bangkok Thailand originalChulalongkorn University, Faculty of Medicine, Department of Medicine, Division of Allergy and Clinical Immunology, Bangkok, Thailand.
                [4 ] orgnameKing Chulalongkorn Memorial Hospital Bangkok Thailand originalKing Chulalongkorn Memorial Hospital, Bangkok, Thailand.
                [5 ] orgnameChulalongkorn University orgdiv1Faculty of Medicine orgdiv2Center of Excellence in Vaccine Research and Development Bangkok Thailand originalChulalongkorn University, Faculty of Medicine, Center of Excellence in Vaccine Research and Development (Chula Vaccine Research Center- Chula VRC), Bangkok, Thailand.
                [6 ] orgdiv2Thai Red Cross Society orgnameKing Chulalongkorn Memorial Hospital orgdiv1Department of Otolaryngology, Head and Neck Surgery Bangkok Thailand originalThai Red Cross Society, King Chulalongkorn Memorial Hospital, Department of Otolaryngology, Head and Neck Surgery, Bangkok, Thailand.
                [7 ] orgnameChulalongkorn University orgdiv1Faculty of Medicine orgdiv2Department of Otolaryngology, Head and Neck Surgery Bangkok Thailand originalChulalongkorn University, Faculty of Medicine, Department of Otolaryngology, Head and Neck Surgery, Bangkok, Thailand.
                [8 ] orgnameMahidol University orgdiv1Faculty of Dentistry orgdiv2Department of Oral Biology Bangkok Thailand originalMahidol University, Faculty of Dentistry, Department of Oral Biology, Bangkok, Thailand.
                Author notes
                Corresponding address: Nakarin Kitkumthorn Department of Oral Biology - Faculty of Dentistry - Mahidol University. Payathai Rd. - Ratchathewi - Bangkok - 10400 - Thailand. Phone: (66)2200-7849 e-mail: nakarinkit@ 123456gmail.com

                Disclosure statement

                The authors declare no conflict of interest.

                Authors' contributions

                Bin-Alee, Fardeela: Data curation (Lead); Formal analysis (Lead); Investigation (Lead); Methodology (Lead); Writing original draft (Lead). Arayataweekul, Areeya: Investigation (Equal); Methodology (Equal); Resources (Lead). Baranaprad, Supranee: Investigation (Lead); Methodology (Equal); Software (Lead). Mahattanasakul, Patnarin: Resources (Lead). Tangjaturonrasme, Napadon: Resources (Equal). Mutirangura, Apiwat: Writing, review & editing (Equal). Kitkumthorn, Nakarin: Conceptualization (Lead); Investigation (Equal); Project administration (Lead); Resources (Equal); Validation (Equal); Writing, review & editing (Lead).

                Author information
                https://orcid.org/0000-0003-0763-341X
                https://orcid.org/0000-0001-6783-2643
                https://orcid.org/0000-0001-9971-4318
                https://orcid.org/0000-0001-6552-5463
                https://orcid.org/0000-0001-5762-703X
                https://orcid.org/0000-0003-0238-942X
                https://orcid.org/0000-0003-0616-6039
                Article
                00467
                10.1590/1678-7757-2020-0124
                7480670
                32901694
                2ba1448d-8aba-4787-8b00-341994468883

                This is an Open Access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

                History
                : 08 May 2020
                : 09 July 2020
                : 14 July 2020
                Page count
                Figures: 6, Tables: 1, Equations: 0, References: 36
                Funding
                Funded by: The Chulalongkorn Academic Advancement into Its 2nd Century
                Award ID: RSA6180078
                Funded by: National Science and Technology Development Agency
                Award ID: FDA-CO-2561-8477-TH
                Categories
                Original Article

                apoptosis,bax/bcl-2,lymphocyte,oral cancer,neoplasm staging
                apoptosis, bax/bcl-2, lymphocyte, oral cancer, neoplasm staging

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