| Human gliomas are the most common tumors in human central nervous system. One of the importmant biological features of gliomas is that local invasion of its constituent neoplastic cells into the surrounding tissue. It is the root cause of high incidence of recurrence in gliomas. Both of the tumor stroma and neoplastic cells corporately contribute to invasion and metastasis of gliomas. It is important for an component of the process in tumor progression that extra cellular matrix and basement membrane are degradated. It is particularly necessary for Matrix metalloproteinases (MMTs) in matrix degradation. In this family, matrix metalloproteinase 2 (MMP-2), tissue specific inhibitors of matrix- metalloproteinases (TIMPs) and membrane-type matrix metalloproteinase 1 (MT1-MMP) have been focused on by many studies. TIMP2 is capable of binding and inhibiting the activity of MMP2. MT1-MMP is a key activator to latent proenzyme of MMP-2. Phosphatase of regenerating liver-3 (PRL-3) is a new member of protein-tyrosine phosphates, which is consistently overexpressed in many metastatic tumor tissues and show close relation to invasion and metastisis. It may be a new therapeutic target in some tumors. Up to now, there is no report on the expression of PRL-3 in gliomas.Experiment 1: MMPs detection and valueObjective To detect the activities and expression of MMPs in gliomas and its value. Methods Freezing tissue blocks and formalin-fixed and paraffin-embedded tissue blocks were retrieved from one hundred and thirty-five glioma cases, which including seventy-six males and fifty-nine females. All cases had no history of the chemotherapy, radiotherapy and immunotherapy. All cases reviewed for the accuracy of diagnoses, according to the criteria of World Health Organization (WHO), all cases were described in four major grades and five tumor types. Seven cases are?grades, fifty-seven cases are?grades, fourtone cases are?grades and thrity cases are?grades. One part of specimens was stored in liquid-nitrogen, the other was formalin-fixed and paraffin-embedded and HE stained. Three cases of normal tissue got from autopsy. Latent and mature proteases of MMP-2 and MMP-9 were detected in one hundred and thirty-five cases of glioma tissues and three normal brain tissues by gelatin zymography. In order to quantitatively analyze the relationship between MMPs and tumor grades , the relationship between activeation of MMP-2 and MT1-MMP was focused on. While, the expressions and correlations of MT1-MMP, MMP-2 were detected by immunohistochemistry and tissue microarray cooperatively. Results 1. There were four or five bands in WHO?and WHO?grades gliomas in gelatin zymography. Molecular weight of MMP-2, intermediate MMP-2, proMMP-2, MMP-9 and proMMP-9 were 62 kD, 64kD, 72kD, 82kD and 92kD respectively. 2. The quantities of MMP-9 in glioma of grade WHO?and WHO?were 75.560±8.380 and 178.170±11.950 A.mm2/g.L respectively, there was a significant difference between them (P<0.05). 3. The quantities of MMP-2 in glioma of grade WHO?and WHO?were 43.586±10.245 and 63.950±6.550 A.mm2/g.L respectively, there was a significant difference between them (P<0.05). 4. Eight cases in glioma of grade WHO?and one case in glioma of grade WHO?showed the intermediate MMP-2 of 64kD, which quantity of grade WHO?glioma was 27.690±19.860 A.mm2/g.L. 5. MT1-MMP immunohistochemically stained in vessel endothelial cells and tumor cells. The positive rates of MT1-MMP protein in normal brain tissue and glioma of grades WHO?,WHO?,WHO?and WHO?were 0,0,15.9%,38.0% and 60.0% respectively. There was a significant difference among glioma grade WHO?, WHO?and WHO?(P<0.05). 6. MMP-2 protein immunohistochemically stained in vessel endothelial cells and glioma tumor cells. The positive rates of MMP-2 protein in normal brain tissue and glioma of grades WHO?,WHO?,WHO?and WHO?were 0,0,18.3%,44.3% and 66.7% respectively. There was a significant difference among tumor grade WHO?,?and?(P<0.05). Conclusion 1. Latent and mature proteases of MMP-2 and MMP-9 can be detected in glioma tissue by gelatin zymography. Mature proteases of MMPs were the key to pathological grades of tumors. 2. In high grade gliomas, the intermediate MMP-2 (MT1-MMP. TIMP-2. MMP-2 complex) was distinguished by gelatin zymography. There was correlation between activation of MMP-2 and the expression of this complex. 3. MT1-MMP and MMP-2 were correlative with tumor angiogenesis and malignant degree, and they corporately contributed to the invasion of gliomas. 4. Hand-worked tissue microarray was successfully used in immunohistochemical staining, which collected lots of valueable experience for using tissue microarray in practice clinical surgical pathology.Experiment 2: PRL-3 detection and valueObjective To detect the PRL-3mRNA and PRL-3 expression in gliomas and its values. Methods One hundred and thirty-five cases of gliomas were studied from the surgical pathology files, including seventy-six males and fifty-nine females. All cases reviewed for the diagnoses according to the current WHO criteria and described in four grades and five types ( pilocytic astrocytoma four cases (grade I), subependymal giant cell astrocytoma three cases (grade I), diffuse astrocytoma fifty-seven cases (grade?), anaplastic astrocytoma forty-one cases (grade?) and glioblastoma thirty cases (grade?). All the examples of cases were divided into two parts, one part was stored in liquid-nitrogen as soon as possible, and other was fixed in buffered-formalin and embedded with paraffin and HE stained. Three normal brain tissues were received from autopsy. Digoxigenin labeled PRL-3 antisense RNA probes were used for in situ hybridization in paraffin embedded glioma tissue slides. The expressions of PRL-3 were detected by immunohistochemistry and tissue microarray . Results No signal of PRL-3 hybridization was observed in tumors of WHO I and normal brain tissues. Signal of PRL-3 mRNA was detected in cytoplasm of glioma cells, especially in gliomas tissues of grade WHO?and WHO?. A positive staining also found in endothelial cells in the tumor tissues which were detected in tumor cells simultaneously. Only little was observed in tumors of WHO?. The positive rates of the expression of PRL-3 protein in normal brain tissue and glioma of WHO?,?,?and?were 0, 0, 5.1%, 23.9% and 50.0% respectively. There was a significant difference between glioma of WHO?,?and?(P<0.05). There was no correlation with other factors such as patient's sex, tumoe site and types of cells in the expression of PRL-3 protein in gliomas (P>0.05). But it was correlative with patient's age (P<0.05). The mean age was 38; the positive rate of cases above the mean age was higher than those below it. The correlation coefficient (r) between MT1-MMP and MMP-2 was 0.425. Conclusion With the increase of tumor grading, the positive rates of signal of PRL-3mRNA and expression of PRL-3 in gliomas increased gradually, which suggested that PRL-3 might play an important role in invasion and progression of gliomas. |
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