Cyclosporine A (CsA) is a widely used immunosuppressant for transplant patients and is also used for the treatment of a wide variety of systemic diseases with immunologic components. A prominent side effect of CsA administration is gingival overgrowth. It has been postulated that CsA alters fibroblast activity through effects on various cytokines such as the interleukins, however, as yet, data concerning the molecular mechanisms involved in connective tissue proliferation are still preliminary in nature. The purpose of this study was to evaluate interleukin‐6 (IL‐6) gene expression in gingival tissues of patients receiving CsA therapy and exhibiting gingival overgrowth. Radioimmunoassay (RIA) demonstrated a significant difference in tissue levels of IL‐6 as mean ± SEM. IL‐6 content in CsA‐stimulated tissue was 184.3 ±30.2 ng/mg total protein versus 23.3 ± 6.5 ng/mg total protein in control tissue. In situ hybridization indicated that overgrown gingival tissues from patients taking CsA had a significantly higher content of IL‐6 mRNA when compared to control tissues. Expressing IL‐6 mRNA levels as silver grains/cell, CsA‐stimulated tissue had 166.9 ± 12.0 grains of IL‐6 mRNA/cell while control tissue had 12.8 ± 3.0 grains of IL‐6 mRNA/cell. These results demonstrate that CsA therapy results in increased levels of IL‐6 protein and IL‐6 mRNA in overgrown human gingival tissues. This is the first report of CsA‐upregulated IL‐6 gene expression in vivo, and may explain in part the molecular mechanisms responsible for CsA‐induced gingival overgrowth. J Periodontol 1994;65:895–903.

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The expression of strictly inducible hsp70 mRNAs and constitutively expressed hsc70 mRNAs was compared in cerebellum and cerebral cortex of control rats, heat-shocked rats, and rats made hyperthermic with amphetamine. An hsc70-specific oligonucleotide probe identified a 2.55-kb mRNA in cerebellum and cerebral cortex of all rats. An hsp70-specific oligonucleotide probe identified a 3.05-kb mRNA and a 3.53-kb mRNA in cerebellum and cerebral cortex of heat-shocked and amphetamine-treated rats, but not in control rats. Quantitation demonstrated that both hsp70 and hsc70 mRNA levels, relative to 18S rRNA levels, were increased following each treatment. The relative levels of both mRNAs were higher in cerebellum than in cerebral cortex. In amphetamine-treated rats, hsc70 mRNA relative levels increased at body temperatures greater than 39 degrees C, whereas hsp70 mRNA synthesis was induced at temperatures greater than 40 degrees C. Total thermal response values and relative levels of both mRNAs were compared. The results suggested that both the transcription and turnover of hsp70 mRNAs differed between cerebellum and cerebral cortex. At equivalent total thermal response values, amphetamine-treated rats had higher relative levels of hsp70 mRNAs than heat-shocked rats, suggesting that amphetamine enhanced the induction of hsp70 mRNAs.

The mechanism by which phenytoin (PHT) induces gingival overgrowth remains unclear. We hypothesized that PHT increases macrophage production of platelet-derived growth factor (PDGF), an important cytokine in connective tissue growth and repair, and that excessive production PDGF in gingiva could lead to redundant growth. To test the hypothesis, rat peritoneal macrophages and human blood monocytes were cultured in the presence of PHT (5 to 20 micrograms/ml medium) or an equal volume of its solvent for 3 days and tested for expression of PDGF-B mRNA by in situ hybridization. Approximately 300 cells/culture well were examined (3 wells/drug level) for positive indication of PDGF-B mRNA. Data were compared by chi square test. All levels of PHT in both cell types induced a 2- to 8-fold increase in PDGF-B mRNA positive cells, significant in all cases at P < 0.001. Northern blot analysis of RNA from similarly cultured rat macrophages confirmed these findings. Cells treated with 10 micrograms PHT/ml medium or solvent revealed 2.2 +/- 0.3 and 1.0 +/- 0.2 (mean +/- SEM) arbitrary units PDGF mRNA respectively (t tests, P < 0.05). Additionally, rat macrophages were cultured in presence of 5 micrograms PHT/medium or its solvent and medium was analyzed for PDGF secretion by radioimmunoassay. Mean values (+/- SEM) were 1.28 +/- 0.49 and 0.78 +/- 0.07 ng/mg protein respectively (t test, P < 0.05). These data showed that PHT augmented the expression of c-sis, the gene for PDGF-B, and offered a possible explanation for PHT-induced gingival overgrowth.

The expression of calbindin-D9K (CaBP9K) and calbindin-D28K (CaBP28K) genes in the reproductive system is well established for rodent and avian species, but not for domestic livestock. This investigation expanded the study of these proteins to include the bovine uterus and examined the levels of CaBP9K and CaBP9K mRNA in the nonpregnant bovine uterus during the estrous cycle. Immunohistochemical studies revealed that CaBP9K was present in all uterine glandular and luminal epithelial cells. In contrast, the closely related calcium binding protein CaBP28K was present in only one to two glandular cells in the samples examined. Neither protein was localized in the myometrium or in the stromal cells of the endometrium. RIA and dot blot hybridization were used to quantify the amount of CaBP9K and CaBP9K mRNA. The levels of both the protein and its mRNA were threefold higher during the luteal phase than during the follicular phase. RIA was also used to determine bovine uterine levels of 17 beta-estradiol and progesterone. Progesterone levels were higher during the luteal phase than during the follicular phase, while 17 beta-estradiol levels were higher during the follicular phase. This investigation represents the first characterization of CaBP9K gene expression in the bovine uterus. It demonstrated that the expression of CaBP9K and CaBP9K mRNA was greatest during the progesterone-dominated luteal phase of the bovine estrous cycle. These results indicated that CaBP9K may be involved in uterine glandular function during the luteal phase.