| Objectives: Oral squamous cell carcinoma(OSCC) is the most common malignant tumor of oral cavity. At present, OSCC is treated with synthetic treatment of operation, radiation and chemotherapy. Therefore, it is necessary to find new methods for treating OSCC. The way through supplying or substituting the deficiency of gene is the new way to treat tumor presently, and it is one of the popular subjects of the medical research fields. Fragile histidine triad(FHIT) gene found resently is a tumor suppressor gene. FHIT is abundantly expressed in normal tissue. Abnormalities of the FHIT gene have been reported in lots of tumor tissue such as head and neck, esophagea, gastric, lung, breast, colorectal carcinoma and so on. At present, the relationship between FHIT and oral squamous cell carcinoma has few reports in the world. There is another major kind of gene system in the genes involved in oncogenesis, besides oncogenes, tumor supressor genes, which is known as mismatch repair(MMR) genes system. The instability of genome which results from defect of MMR genes is thought as a new oncogenesis way. The symbol of genome instabillity is microsatellite instability(MSI) and loss of protein. hMLH1 and hMSH2 are two important gene of MMR. Recent investigations demonstrated FHIT inactivation may be a consequence of defects in mismatch repair genes. Then are there among them correlations in the carcinogenesis and progresssion of OSCC? It has not been reported presently. We adopted immunohistochemical method in the study to investigate the expression, significance and correlation of FHIT hMLH1 and hMSH2 in oral squamous cell carcinomas(OSCC), in order to clarify the mechanism of FHIT, hMSH2and hMLH1 in OSCC, then to provide theoretical evidence for preventing and treating OSCC in clinic. Methods: Paraffin-embedded blocks of 69 OSCC tissues and 40 normal oral tissues were selected and divided into experimental group (OSCC) and control group (normal mucosa), respectively. 5 sections with thickness of 4-5μm were cut from each block, one for HE staining, one for negative control, the others for immunohistochemical staining of FHIT,hMSH2 and hMLH1 respectively. The immunohistochemical staining was operated as following: the paraffin sections were deparaffinized by normal method; placed them in methanol containing 3% H2O2 for 25 min to block endogenous peroxidase activity; put them into microwave oven to repair antigens for 15 min at 92-98℃, cooled at room temperature; the section were treated with normal goats serum for 30min at 37℃; after wipping the serum off, the sections were incubated with the primary antibodies forone night at 4℃; the sections were treated with biotinglated secondary antibodies for 25 min at 37 ℃; and with peroxidaze-conjugated streptavidin for 20 min at 37℃; stained with DAB and conterstained with hematoxylin, dehydrated in graded rthamal, cleared in xylene and finally mounted. Except for the normal goat serum-blocking step, the sections were washed with PBS for 3×5 min after each treatment. Positive breast tissues expressing FHIT, positive colorectal tissues expressing hMSH2 and hMLH1 were used as positive control, respectively . As a negative control, the sections were treated with PBS instead of the primary antibodies. Specific staining was identified by the presence of yellow or brown reaction products in cytoplasm and/or nucleus. A semiquantitative scoring system was used for FHIT as follows: for each tissue specimen the rate of cell stain and intensity of staining with FHIT antibody is graded on a score of 1—3 to assign the score respectively. The intensity of staining was recorded as: absent/weak, 1; moderate, 2; and strong, 3. The extent of immunostaining was scored based on the percentage of positive cells: 10%, 1; 10-50%, 2; and >50%, 3. The two scores were then multiplied to give a composite score (1-9) for each tumor. Composite scores of 1-3 were defined as marked reduction or absence of FHIT protein expression, while scores >3 were considered positive for FHIT. It is used for hMSH2 and hMLH1 as follows: The presence of staining in >5% of cells was indicated as(+),while <5% was indicated as(-).The data is analyzed by Chi-square test and Spearman rank correlation. Results 1. In normal mucosas, FHIT mostly expressd in the basal and spinosum, few expressed in the granulosum. Positive cells are dense and strong staining in the basal epithelium. FHIT expressd both in nuclear and cytoplasm of cells . In OSCC, FHIT mostly expressd in cytoplasm. Positive rates of FHIT in OSCC and controls were 46.4% and 77.5% respectively. These differences between OSCC and normal groups were statistically significant (P<0.05) . 2. In normal mucosas, hMSH2 mostly expressd in nuclear of cells in the basal and spinosum. Some cases expressed in cytoplasm simultaneously, brown or yellow reaction products. hMSH2 mostly expressd in nuclear of cells in OSCC, few cases in cytoplasm, yellow reaction products. Positive rates of hMSH2 in OSCC and controls were 55.1% and 80.0% respectively. These differences between OSCC and normal groups were statistically significant (P<0.05). In OSCC, no correlation had been found between FHIT and hMSH2(P>0.05). 3. In normal mucosas, hMLH1 mostly expressd in cytoplasm of cells, some cases in nuclear simultaneously, brown or yellow reaction products. Positive cells are dense and strong staining in the basal epithelium. hMLH1 mostly expressd in cytoplasm of cells in OSCC, few cases in nuclear. Positive rates of FHIT in OSCC and control were 47.8% and 77.5% respectively. These...
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