Periodontal disease in adults with untreated congenital growth hormone deficiency: a case-control study

P>Aim The aim of this study was to investigate the possible associations between isolated growth hormone deficiency (IGHD) and periodontal attachment loss (PAL) in adults affected by congenital IGHD. Materials and methods Forty-five previously identified IGHD subjects were eligible for this study. The final study sample comprised 32 cases (gender:20M/12F; age:44.8 +/- 17.5) matched for age, gender, diabetes, smoking status and income to 32 controls (non-IGHD subjects). Participants were submitted to a full-mouth clinical examination of six sites per tooth and were interviewed using a structured, written questionnaire. Periodontitis was defined as proximal PAL >= 5 mm affecting >= 30% of teeth. Results No significant differences were observed in the percentage of sites with visible plaque between IGHD and non-IGHD subjects (59.4% versus 46.9%, p=0.32). IGHD subjects had significant less supragingival calculus (31.3% versus 59.4%, p=0.02) and more bleeding on probing (71.9% versus 18.8%, p < 0.01) than controls. PAL >= 5 mm was significantly more prevalent (100% versus 71.9%, p < 0.01) and affected more teeth (30.5% versus 6.7%, p < 0.01) in cases than in controls. After adjusting for supragingival calculus, IGHD cases had a higher likelihood of having periodontitis than controls (OR=17.4-17.8, 95% CI=2.3-134.9, p=0.004-0.005). Conclusion Congenital IGHD subjects have a greater chance of having PAL.

Growth hormone is permissive for skeletal adaptation to mechanical loading

The Lewis dwarf (DW) rat was used as a model to test the hypothesis that growth hormone (GH) is permissive for new bone formation induced by mechanical loading in vivo. Adult female Lewis DW rats aged 6.2 +/- 0.1 months (187 +/- 18 g) were allocated to four vehicle groups (DW), four GH treatment groups at 32.5 mug/100 g body mass (DWGH1), and four GH treatment groups at 65 mug/100 g (DWGH2). Saline vehicle or GH was injected intraperitoneally (ip) at 6:30 p.m. and 6:30 a.m. before mechanical loading of tibias at 7:30 a.m. A single period of 300 cycles of four-point bending was applied to right tibias at 2.0 Hz, and magnitudes of 24, 29, 38, or 48N were applied. Separate strain gauge analyses in 5 DW rats validated the selection of loading magnitudes. After loading, double-label histomorphometry was used to assess bone formation at the periosteal surface (Ps.S) and endocortical surface (Ec.S) of tibias. Comparing left (unloaded) tibias among groups, GH treatment had no effect on bone formation. Bone formation in tibias in DW rats was insensitive to mechanical loading. At the Ec.S, mechanically induced lamellar bone formation increased in the DWGH2 group loaded at 48N (p < 0.05), and no significant increases in bone formation were observed among other groups. The percentage of tibias expressing woven bone formation (Wo.B) at the Ps.S was significantly greater in the DWGH groups compared with controls (p < 0.05). We concluded that GH influences loading-related bone formation in a permissive manner and modulates the responsiveness of bone tissue to mechanical stimuli by changing thresholds for bone formation.

Influence of growth hormone on the mandibular condylar cartilage of rats

Growth hormone (GH) stimulates mandibular growth but its effect on the mandibular condylar cartilage is not well. understood. Objective: This study was designed to understand the influence of GH on mitotic activity and on chondrocytes maturation. The effect of GH on cartilage thickness was also determined. Design: An animal model witt differences in GH status was determined by comparing mutant Lewis dwarf rats with reduced pituitary GH synthesis (dwarf), with normal rats and dwarf animals treated with GH. Six dwarf rats were injected with GH for 6 days, while other six normal rats and six dwarf rats composed other two groups. Mandibular condylar tissues were processed and stained for Herovici's stain and immunohistochemistry, for proliferating cell nuclear antigen (PCNA) and alkaline phosphatase (ALP). Measurements of cartilage thickness as well as the numbers of immunopositive cells for each antibody were analysed by one-way analysis of variance. Results: Cartilage thickness was significantly reduced in the dwarf animals treated with GH. PCNA expression was significant lower in the dwarf rats, but significantly increased when these animals were treated with GH. ALP expression was significant higher in the dwarf animals, while it was significantly reduced in the dwarf animals treated with GH. Conclusions: The results from this study showed that GH stimulates mitotic activity and delays cartilage cells maturation in the mandibular condyte. This effect at the cellular Level may produce changes in the cartilage thickness. (C) 2004 Elsevier Ltd. All rights reserved.

Growth hormone induces bone morphogenetic proteins and bone-related proteins in the developing rat periodontium

The hypothesis that growth hormone (GH) up-regulates the expression of enzymes, matrix proteins, and differentiation markers involved in mineralization of tooth and bone matrices was tested by the treatment of Lewis dwarf rats with GH over 5 days, The molar teeth and associated alveolar bone were processed for immunohistochemical demonstration of bone morphogenetic proteins 2 and 4 (BMP-2 and -4), bone morphogenetic protein type IA receptor (BMPR-IA), bone alkaline phosphatase (ALP), osteocalcin (OC), osteopontin (OPN), bone sialoprotein (BSP), and E11 protein (E11), The cementoblasts, osteoblasts, and periodontal ligament (PDL) cells responded to GH by expressing BMP-2 and -4, BMPR-IA, ALP, OC, and OPN and increasing the numbers of these cells. No changes were found in patterns of expression of the late differentiation markers BSP and E11 in response to GH, Thus, GH evokes expression of bone markers of early differentiation in cementoblasts, PDL cells, and osteoblasts of the periodontium. We propose that the induction of BMP-2 and -4 and their receptor by GH compliments the role of GH-induced insulin-like growth factor 1 (IGF-1) in promoting bone and tooth root formation.

Dietary intervention with green dwarf banana flour (Musa sp AAA) prevents intestinal inflammation in a trinitrobenzenesulfonic acid model of rat colitis

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

T-cell receptor (TCR) usage in Lewis rat experimental autoimmune encephalomyelitis: TCR beta-chain-variable-region V beta 8.2-positive T cells are not essential for induction and course of disease.

Predominant usage of V beta 8.2 gene segments, encoding a T-cell receptor (TCR) beta chain variable region, has been reported for pathogenic Lewis rat T cells reactive to myelin basic protein (MBP). However, up to 75% of the alpha/beta T cells in a panel of MBP-specific T-cell lines did not display TCR V beta 8.2, V beta 8.5, V beta 10, or V beta 16 elements. To further investigate TCR usage, we sorted the T-cell lines for V beta 8.2- and V beta 10-positive T cells or depleted the lines of cells with these TCRs. V beta 8.2-positive T cells and one of the depleted T-cell lines strongly reacted against the MBP peptide MBP-(68-88). The depleted T-cell line caused marked experimental autoimmune encephalomyelitis (EAE) even in Lewis rats in which endogenous V beta 8.2-positive T cells had been eliminated by neonatal treatment with anti-V beta 8.2 monoclonal antibodies. T-cell hybridomas generated from this line predominantly used V beta 3 TCR genes coexpressed with TCR V alpha 2 transcripts, which were also used by V beta 8.2-positive T cells. Furthermore, V beta 10-positive T cells reactive to MBP-(44-67) were encephalitogenic when injected immediately after positive selection. After induction of EAE by sorted V beta 8.2- or V beta 10-positive T-cell lines, immunocytochemical analysis of the spinal cord tissue showed a predominance of the injected TCR or of nontypable alpha/beta T cells after injection of the depleted line. Our results demonstrate heterogeneity of TCR beta-chain usage even for a single autoantigen in an inbred strain. Moreover, V beta 8.2-positive T cells are not essential for the induction and progression of adoptive-transfer EAE.

Rat Adipose Tissue-Derived Stem Cells Transplantation Attenuates Cardiac Dysfunction Post Infarction and Biopolymers Enhance Cell Retention

Background: Cardiac cell transplantation is compromised by low cell retention and poor graft viability. Here, the effects of co-injecting adipose tissue-derived stem cells (ASCs) with biopolymers on cell cardiac retention, ventricular morphometry and performance were evaluated in a rat model of myocardial infarction (MI). Methodology/Principal Findings: (99m)Tc-labeled ASCs (1 x 10(6) cells) isolated from isogenic Lewis rats were injected 24 hours post-MI using fibrin a, collagen (ASC/C), or culture medium (ASC/M) as vehicle, and cell body distribution was assessed 24 hours later by gamma-emission counting of harvested organs. ASC/F and ASC/C groups retained significantly more cells in the myocardium than ASC/M (13.8+/-2.0 and 26.8+/-2.4% vs. 4.8+/-0.7%, respectively). Then, morphometric and direct cardiac functional parameters were evaluated 4 weeks post-MI cell injection. Left ventricle (LV) perimeter and percentage of interstitial collagen in the spare myocardium were significantly attenuated in all ASC-treated groups compared to the non-treated (NT) and control groups (culture medium, fibrin, or collagen alone). Direct hemodynamic assessment under pharmacological stress showed that stroke volume (SV) and left ventricle end-diastolic pressure were preserved in ASC-treated groups regardless of the vehicle used to deliver ASCs. Stroke work (SW), a global index of cardiac function, improved in ASC/M while it normalized when biopolymers were co-injected with ASCs. A positive correlation was observed between cardiac ASCs retention and preservation of SV and improvement in SW post-MI under hemodynamic stress. Conclusions: We provided direct evidence that intramyocardial injection of ASCs mitigates the negative cardiac remodeling and preserves ventricular function post-MI in rats and these beneficial effects can be further enhanced by administrating co-injection of ASCs with biopolymers.

A comparison of lipopolysaccharide-induced hepatitis in ethanol-fed Wistar and Lewis rats

Elevated concentrations of plasma proinflammatory cytokines have been detected in patients with alcoholic hepatitis (AH) and in a model of lipopolysaccharide-induced hepatitis in ethanol-fed Wistar rats. These cytokines have been implicated in the pathogenesis of the liver damage. Considering the likely involvement of the immune system in AH, and the frequent use of Lewis rats in autoimmune disease models, Lewis rats were examined in the model to determine whether they would more closely mimic the immune status of a chronic alcoholic and be a preferable strain for use in future experiments. Lipopolysaccharide-induced hepatic tumor necrosis factor-cu, interleukin-1 alpha, interleukin-1 beta, and interleukin-6 mRNA expression was examined in both rat strains. The overall pattern of histological (panlobular piecemeal necrosis) and biochemical liver damage (plasma ALT levels), and cytokine expression was similar in both strains. Thus, it would appear that, despite the known susceptibility of Lewis rats to autoimmune phenomena, they do not respond to the experimental regime significantly better than Wistar rats. This study confirms that unknown mediators are contributing to the liver damage seen in this model and possibly in AH.

Histologic and immunohistochemical responses after aortic valve allografts in the rat

Background. Human aortic valve allografts elicit a cellular and humoral immune response. It is not clear whether this is important in promoting valve damage. We investigated the changes in morphology, cell populations, and major histocompatibility complex antigen distribution in the rat aortic valve allograft. Methods. Fresh heart valves from Lewis rats were transplanted into the abdominal aorta of DA rats. Valves from allografted, isografted, and presensitized recipient rats were examined serially with standard morphologic and immunohistochemical techniques. Results. In comparison with isografts, the allografts were infiltrated and thickened by increased numbers of CD4(+) and CD8(+) lymphocytes, macrophages, and fibroblasts. Thickening of the valve wall and leaflet and the density of the cellular infiltrate was particularly evident after presensitization. Endothelial cells were frequently absent in presensitized allografts whereas isografts had intact endothelium. Cellular major histocompatibility complex class I and II antigens in the allograft were substantially increased. A long-term allograft showed dense fibrosis and disruption of the media with scattered persisting donor cells. Conclusions. The changes in these aortic valve allograft experiments are consistent with an allograft immune response and confirm that the response can damage aortic valve allograft tissue. (C) 1998 by The Society of Thoracic Surgeons.

EIU in the rat promotes the potential of syngeneic retinal cells injected into the vitreous cavity to induce PVR.

PURPOSE: To determine whether syngeneic retinal cells injected in the vitreous cavity of the rat are able to initiate a proliferative process and whether the ocular inflammation induced in rats by lipopolysaccharide (LPS) promotes this proliferative vitreoretinopathy (PVR). METHODS: Primary cultured differentiated retinal Müller glial (RMG) and retinal pigmented epithelial (RPE) cells isolated from 8 to 12 postnatal Lewis rats were injected into the vitreous cavity of 8- to 10-week-old Lewis rats (10(5) cells/eye in 2 microlieter sterile saline), with or without the systemic injection of 150 microgram LPS to cause endotoxin-induced uveitis (EIU). Control groups received an intravitreal injection of 2 microliter saline. At 5, 15, and 28 days after cell injections, PVR was clinically quantified, and immunohistochemistry for OX42, ED1, vimentin (VIM), glial fibrillary acidic protein (GFAP), and cytokeratin was performed. RESULTS: The injection of RMG cells, alone or in combination with RPE cells, induced the preretinal proliferation of a GFAP-positive tissue, that was enhanced by the systemic injection of LPS. Indeed, when EIU was induced at the time of RMG cell injection into the vitreous cavity, the proliferation led to retinal folds and localized tractional detachments. In contrast, PVR enhanced the infiltration of inflammatory cells in the anterior segment of the eye. CONCLUSIONS: In the rat, syngeneic retinal cells of glial origin induce PVR that is enhanced by the coinduction of EIU. In return, vitreoretinal glial proliferation enhanced the intensity and duration of EIU.

The effect of COX-2 inhibitors on the formation of heterotopic bone in a rat model : O1323

Aims: 1) to create a new and reproducible animal model to produce heterotopic ossification (HO) 2) to be able to exactly quantify the amount of HO using a microCT scan and 3) to prove the hypothesis that COX-2 inhibitors are efficacious in the prevention of HO. Methods: We developed a IACUC-approved Lewis rat model, in which the ventral side of the right femur was scraped to mechanically disrupt the periosteum. By clamping the vastus intermedius ischemic injury to the muscle was produced to enhance HO. Finally homologous bone marrow from a donor rat was placed on the anterior surface of the femur. Half of the study group (8 rats) received chow mixed with a COX-2 inhibitor, while the other half received normal chow. After 6 weeks the animals were sacrificed, the femurs removed and imaged by microCT. Grading of HO was based on the thickness of ectopic bone as evaluated in a blinded fashion by 3 independent observers. Results: All animals developed bilateral HO. Rats treated with COX-2 inhibitors developed significantly less ectopic bone than the control group rats. Conclusions: The results suggest that we have created a very reliable, reproducible model to form ectopic bone in rats. Using the microCT we can precisely quantify the amount of HO. We have been able to show that COX-2 inhibitors significantly decrease the amount of HO formation and are thus a good alternative to non-specific NSAIDs with their potential serious side effects on the gastrointestinal tract and on hemo-stastis.

Anti-vascular endothelial growth factor acts on retinal microglia/macrophage activation in a rat model of ocular inflammation.

PURPOSE: To evaluate whether anti-vascular endothelial growth factor (VEGF) neutralizing antibodies injected in the vitreous of rat eyes influence retinal microglia and macrophage activation. To dissociate the effect of anti-VEGF on microglia and macrophages subsequent to its antiangiogenic effect, we chose a model of acute intraocular inflammation. METHODS: Lewis rats were challenged with systemic lipopolysaccharide (LPS) injection and concomitantly received 5 µl of rat anti-VEGF-neutralizing antibody (1.5 mg/ml) in the vitreous. Rat immunoglobulin G (IgG) isotype was used as the control. The effect of anti-VEGF was evaluated at 24 and 48 h clinically (uveitis scores), biologically (cytokine multiplex analysis in ocular media), and histologically (inflammatory cell counts on eye sections). Microglia and macrophages were immunodetected with ionized calcium-binding adaptor molecule 1 (IBA1) staining and counted based on their differential shapes (round amoeboid or ramified dendritiform) on sections and flatmounted retinas using confocal imaging and automatic quantification. Activation of microglia was also evaluated with inducible nitric oxide synthase (iNOS) and IBA1 coimmunostaining. Coimmunolocalization of VEGF receptor 1 and 2 (VEGF-R1 and R2) with IBA1 was performed on eye sections with or without anti-VEGF treatment. RESULTS: Neutralizing rat anti-VEGF antibodies significantly decreased ocular VEGF levels but did not decrease the endotoxin-induced uveitis (EIU) clinical score or the number of infiltrating cells and cytokines in ocular media (interleukin [IL]-1β, IL-6, tumor necrosis factor [TNF]-α, and monocyte chemoattractant protein [MCP]-1). Eyes treated with anti-VEGF showed a significantly decreased number of activated microglia and macrophages in the retina and the choroid and decreased iNOS-positive microglia. IBA1-positive cells expressed VEGF-R1 and R2 in the inflamed retina. CONCLUSIONS: Microglia and macrophages expressed VEGF receptors, and intravitreous anti-VEGF influenced the microglia and macrophage activation state. Taking into account that anti-VEGF drugs are repeatedly injected in the vitreous of patients with retinal diseases, part of their effects could result from unsuspected modulation of the microglia activation state. This should be further studied in other ocular pathogenic conditions and human pathology.