Guilherme Negrão de Souza

Alterações histoquímicas das glicosaminoglicanas na cérvice uterina no final da prenhez da rata albina após ministração local de hialuronidase

PURPOSE: to study the histochemical changes related to the uterine cervix glycosaminoglycan of the albino female rat, after local ministration of hyaluronidases at the end of pregnancy. METHODS: ten female rats with positive pregnancy tests were randomly distributed in two numerically equal groups. The control group (Cg) was built up with rats that received a single dose of 1 mL of distilled water in the uterine cervix, under anesthesia, at the 18th pregnancy day. In the experimental group (Exg), the rats received 0.02 mL of hyaluronidase, diluted in 0.98 mL of distilled water (1 mL as a total), under the same conditions as the Cg. At the 20th pregnancy day, the rats were anesthetized once again and submitted to dissection, and the cervix prepared for histochemical study with alcian blue dye and its blockades (pH=0.5, pH=2.5, methylation and saponification). RESULTS: strongly positive reaction in the lamina propria (+3) has been seen in the Cg, and negative reaction in the Exg, with pH=0.5 alcian blue staining. With pH=2.5, staining has also been strongly positive (+4) in the Cg, and weakly positive (+1) in the Exg slide. After methylation, both groups have shown negative reaction, with pH=2.5 alcian blue staining. The lamina propria staining became negative after methylation in both groups, followed by saponification and enzymatic digestion on slide. CONCLUSIONS: there is clear predominance of sulphated glycosaminoglycans in the Cg as compared to the Exg and a small amount of identified carboxylated glycosaminoglycans in the Exg. The changes evidenced in the extracellular matrix have suggested that the hyaluronidase injected in the uterine cervix has promoted biochemical changes compatible with cervix maturation.

The expression of glycosaminoglycans and proteoglycans in the uterine cervix of albino rats after local hyaluronidase infusion

Abstract Objective: To assess the local effect of hyaluronidase injection on the expression of glycosaminoglycans (GAGs) and proteoglycans (PGs) in the extracellular matrix of the uterine cervix from pregnant albino rats. Methods: Ten pregnant rats were divided into two groups on day 18 of pregnancy. The experimental group (Gexp) of rats received an intracervical infusion of 0.02 mL of hyaluronidase diluted to 1 mL with distilled water, whereas the control group (Gc) received 1 mL of distilled water. On day 20 of pregnancy, the pregnant rats were sacrificed and the uterine cervixes from all rats were then dissected. The qualitative expression of hyaluronic acid (HA) was assessed by immunohistochemistry and quantified by sandwich ELISA. To compare the quantitative GAG values between groups, a Student’s t-test for independent samples was performed. PGs were also assessed by immunohistochemical analysis. Results: The electrophoretic profile of newly synthesized radioactively labeled GAGs degraded by specific enzymes showed that there were two predominant GAGs in both Gc and Gexp, i.e. heparan sulfate (HS) and a mixture of hondroitin sulfate (CS) and dermatan sulfate (DS). The concentrations of GAGs showed a significant reduction of CS/DS (p < 0.004) and HS (p < 0.005) relative to Gc. HA staining was less intense in the lamina propria and area surrounding the blood vessels in Gexp compared to Gc. The HA contents were also significantly reduced (p < 0.012). Conclusions: Intracervical hyaluronidase infusion promoted a significant reduction in the concentration of sulfated GAGs as assessed by both qualitative (histochemical) and quantitative (fluorometric) measurements of HA.

Alterações morfológicas e na contagem de fibras colágenas induzidas pela hialuronidase no colo de ratas prenhes

PURPOSE: to assess the morphological and morphometric alterations in the uterine cervix of pregnant albino rats determined by local hyaluronidase administration. METHODS: ten rats with a positive pregnancy test were randomly distributed into two equal groups. The control group consisted of rats that received a single dose of 1 mL distilled water in the uterine cervix, on gestational day 18, under anesthesia. The experimental group consisted of rats that received 0.02 mL hyaluronidase, diluted in 0.98 ml distilled water (total = 1 mL), in the same conditions as those of the control group. On day 20, the rats were anesthetized and submitted to dissection. The uterine cervix was prepared for morphological and morphometric study at light microscopy (hematoxylin and eosin, and Masson trichrome). RESULTS: in the experimental group, greater thinning of the superficial mucified epithelium was observed, with lamina propria rich in blood vessels and eosinophils. Diversely, the control group showed a large concentration of collagen fibers. The histometric analysis in the experimental group was characterized by a smaller number of collagen fibers (mean = 248 versus 552 of control; SD = 49.7 versus 31.1 of control). The parametric method (Students t test) showed a significant difference between groups (p<0.0001). CONCLUSION: the local use of hyaluronidase in the cervix of pregnant rats determined predominance of loose connective tissue and a smaller concentration of collagen fibers.