Skeletal Effects of a Prolonged Oral Metformin Treatment in Adult Wistar Rats.

INTRODUCTION: We previously showed that a 3-week oral metformin (MET) treatment enhances the osteogenic potential of bone marrow stromal cells (BMSCs) and improves several bone histomorphometric parameters in Wistar rats with metabolic syndrome (MetS). However, the skeletal effects of extended periods of MET need to be completely elucidated. Hence, in this study, the impact of a prolonged (3-month) MET treatment was investigated on bone architecture, histomorphometric and biomechanics variables, and osteogenic potential of BMSCs in Wistar rats with or without MetS. MATERIALS AND METHODS: Young male Wistar rats (n=36) were randomized into four groups (n=9) that received either 20% fructose (F), MET (MET), F plus MET treatments (FMET), or drinking water alone (Veh). Rats were euthanized, blood was collected, and bones were dissected and processed for peripheral quantitative computed tomography (pQCT) analysis, static and dynamic histomorphometry, and bone biomechanics. In addition, BMSCs were isolated to determine their osteogenic potential. RESULTS: MET affected trabecular and cortical bone, altering bone architecture and biomechanics. Furthermore, MET increased the pro-resorptive profile of BMSCs. In addition, fructose-induced MetS practically did not affect the the structural or mechanical variables of the skeleton. CONCLUSION: A 3-month treatment with MET (with or without MetS) affects bone architecture and biomechanical variables in Wistar rats.

Femoral Structure and Biomechanical Characteristics in Sanfilippo Syndrome Type-B Mice.

Sanfilippo syndrome Type-B, also known as mucopolysaccharidosis IIIB (MPS IIIB), accounts for approximately one-third of all Sanfilippo syndrome patients and is characterized by a similar natural history as Type-A. Patients suffer from developmental regression, bone malformation, organomegaly, GI distress, and profound neurological deficits. Despite human trials of enzyme replacement therapy (ERT) (SBC-103, AX250) in MPS IIIB, there is currently no FDA approved treatment and a few palliative options. The major concerns of ERT and gene therapy for the treatment of bone malformation are the inadequate biodistribution of the missing enzyme, N-acetyl-α-glucosaminidase (NAGLU), and that the skeleton is a poorly hit target tissue in ERT and gene therapy. Each of the four known human types of MPS III (A, B, C, and D) is usually regarded as having mild bone manifestations, yet it remains poorly characterized. This study aimed to determine bone mineral content (BMC), volumetric bone mineral density (vBMD), and biomechanical properties in femurs MPS IIIB C57BL/6 mice compared to phenotypic control C57BL/6 mice. Significant differences were observed in MPS IIIB mice within various cortical and cancellous bone parameters for both males and females (p < 0.05). Here, we establish some osteogenic manifestations of MPS IIIB within the mouse model by radiographic and biomechanical tests, which are also differentially affected by age and sex. This suggests that some skeletal features of the MPS IIIB mouse model may be used as biomarkers of peripheral disease correction for preclinical treatment of MPS IIIB.

Bone structural, biomechanical, and histomorphometric characteristics of the hindlimb skeleton in the marsh rice rat (Oryzomys palustris).

The rice rat (Oryzomys palustris) is a nonconventional laboratory rodent species used to model some human bone disorders. However, no studies have been conducted to characterize the postcranial skeleton. Therefore, we aimed to investigate age- and gender-related features of the hindlimb skeleton of this species. We used femurs and tibiae from 94 rats of both genders aged 4-28 weeks. Bone mineral content (BMC), volumetric bone mineral density (vBMD), and biomechanical properties were determined in femurs. In addition, bone histomorphometry of tibiae was conducted to assess bone cell activities and bone turnover over time. Bone length, total metaphysis BMC and vBMD, mid-diaphyseal BMC and vBMD, cortical thickness, and cortical area progressively augmented with age. Whereas the increase in these parameters plateaued at age 16-22 weeks in female rats, they continued to rise to age 28 weeks in male rats. Furthermore, bone strength parameters increased with age, with few differences between genders. We also observed a rapid decrease in longitudinal growth between age 4 and 16 weeks. Whereas young rats had a greater bone formation rate and bone turnover, older rice rats had greater bone volume and trabecular thickness, with no differences between genders. (a) Sexual dimorphism in the rice rat becomes grossly evident at age 16 weeks; (b) the age-related increases in bone mass, structural cortical parameters, and in some biomechanical property parameters plateau at an older age in male than in female rats; and (c) bone growth and remodeling significantly decreased with age irrespective of the gender.

Diet-induced Generalized Periodontitis in Lewis Rats.

Periodontitis is an important public health concern worldwide. Because rodents from the genus Rattus are resistant to spontaneous periodontitis, experimental periodontitis must be initiated by mechanical procedures and interventions. Due to their exacerbated Th1 response and imbalanced Th17 regulatory T-cell responses, Lewis rats are highly susceptible to inducible inflammatory and autoimmune diseases. We hypothesized that feeding Lewis rats a diet high in sucrose and casein (HSC) would alter the oral microenvironment and induce inflammation and the development of periodontitis lesions without mechanical intervention. A baseline group (BSL, n = 8) was euthanized at age 6 wk. Beginning at 6 wk of age, 2 groups of Lewis rats were fed standard (STD, n = 12) or HSC (n = 20) chow and euthanized at 29 wk of age. We evaluated the degree of periodontitis through histology and µCT of maxillae and mandibles. The HSC-induced inflammatory response of periodontal tissues was assessed by using immunohistochemistry. Gene expression analysis of inflammatory cytokines associated with Th1 and Th17 responses, innate immunity cytokines, and tissue damage in response to bacteria were assessed also. The potential systemic effects of HSC diet were evaluated by assessing body composition and bone densitometry endpoints; serum leptin and insulin concentrations; and gene expression of inflammatory cytokines in the liver. Placing Lewis rats on HSC diet for 24 wk induced a host Th1-immune response in periodontal tissues and mild to moderate, generalized periodontitis characterized by inflammatory cell infiltration (predominantly T cells and macrophages), osteoclast resorption of alveolar bone, and hyperplasia and migration of the gingival epithelium. HSC-fed Lewis rats developed periodontitis without mechanical intervention in the oral cavity and in the absence of any noteworthy metabolic abnormalities. Consequently, the rat model we described here may be a promising approach for modeling mild to moderate periodontitis that is similar in presentation to the human disease.

Zoledronate treatment duration is linked to bisphosphonate-related osteonecrosis of the jaw prevalence in rice rats with generalized periodontitis.

OBJECTIVES: To determine the extent that zoledronate (ZOL) dose and duration is associated with bisphosphonate-related osteonecrosis of the jaw (BRONJ) prevalence in rice rats with generalized periodontitis (PD), characterize structural and tissue-level features of BRONJ-like lesions in this model, and examine the specific anti-resorptive role of ZOL in BRONJ. MATERIALS AND METHODS: Rice rats (n = 228) consumed high sucrose-casein diet to enhance generalized PD. Groups of rats received 0, 8, 20, 50 or 125 µg/kg IV ZOL/4 weeks encompassing osteoporosis and oncology ZOL doses. Rats from each dose group (n = 9-16) were necropsied after 12, 18, 24 and 30 weeks of treatment. BRONJ-like lesion prevalence and tissue-level features were assessed grossly, histopathologically and by MicroCT. ZOL bone turnover effects were assessed by femoral peripheral quantitative computed tomography, serum bone turnover marker ELISAs and osteoclast immunolabelling. RESULTS: Prevalence of BRONJ-like lesions was significantly associated with (a) ZOL treatment duration, but plateaued at the lowest oncologic dose, and (b) there was a similar dose-related plateau in the systemic anti-resorptive effect of ZOL. ZOL and BRONJ-like lesions also altered the structural and tissue-level features of the jaw. CONCLUSION: The relationship between BRONJ-like lesion prevalence and ZOL dose and duration varies depending on the co- or pre-existing oral risk factor. At clinically relevant doses of ZOL, BRONJ-like lesions are associated with anti-resorptive activity.

Comparison of Isoflurane, Ketamine-Dexmedetomidine, and Ketamine-Xylazine for General Anesthesia during Oral Procedures in Rice Rats (Oryzomys palustris).

Rice rats (Oryzomys palustris) are an unconventional laboratory species that has been used to study photoperiodicity, periodontitis, and osteonecrosis of the jaw. Interventional procedures that require anesthesia, including oral procedures, are sometimes necessary in preclinical settings. The use of anesthetics including isoflurane and ketamine combined with α2-adrenoreceptor agonists, such as dexmedetomidine and xylazine, is well-established for laboratory rodents. However, their effects have been studied only modestly in rice rats. The aims of this study were to 1) determine the safety and consistency of 3 common anesthetic modalities in rice rats; 2) compare the physiologic and clinical responses to these anesthetics, and 3) verify the effectiveness of the most successful modality by testing it during an oral procedure (tooth extraction). Isoflurane, intraperitoneal ketamine-dexmedetomidine, and intraperitoneal ketamine-xylazine were evaluated by using a crossover design, in which each rat received all of the anesthetics. Compared with ketamine-dexmedetomidine and ketamine-xylazine, isoflurane inhalation through a nose cone produced more rapid induction, entry to a surgical plane of anesthesia, and initial recovery. In addition, isoflurane produced optimal anesthesia throughout the procedure for most rats. Unlike ketamine-dexmedetomidine and ketamine-xylazine, isoflurane did not alter rectal temperature, SpO2, or respiratory rate during the surgical tolerance period, whereas ketamine-dexmedetomidine and ketamine-xylazine decreased rectal temperature during the last stage of anesthesia and induced cardiorespiratory depression. Furthermore, 2 rats experienced negative outcomes warranting euthanasia: one after receiving ketamine-dexmedetomidine, and the other after ketamine-xylazine anesthesia. In conclusion, isoflurane was the most reliable and effective anesthetic in rice rats and maintained a surgical depth of anesthesia for as long as 30 min, thus supporting successful tooth extractions.

Prevalence of Food Impaction-Induced Periodontitis in Conventionally Housed Marsh Rice Rats (Oryzomys palustris).

Marsh rice rats (Oryzomys palustris) fed a pelleted diet high in sucrose and casein have been used as a model for moderate to severe periodontitis. Here we characterize the prevalence, location, and histopathologic features of food-impaction lesions (FIL), a unique type of oral event, in rice rats fed standard pelleted rodent chow from weaning until 34 wk of age. Healthy female rats (n = 90; age, 4 wk) were weaned into groups (n = 10 to 24) and were euthanized at 4, 16, 22, 28, or 34 wk of age. At necropsy, high-resolution photographs of the 4 jaw quadrants were examined by 3 independent observers to determine the presence, number, and location of FIL. In addition, gross periodontitis was scored (scale, 0 to 4), and the hemimaxillar surface area containing FIL was measured. Serial sections of decalcified jaws were assessed histologically. The prevalence of FIL increased with age, and was 0% (baseline), 59.1%, 69.6%, 81.8% and 80.0% in rats at age 4, 16, 22, 28, and 34 wk, respectively. FIL were predominantly located (93.9%) in the maxillary palatal surfaces of the interproximal area between molars 2 and 3 and did not affect mandibular surfaces. The percentage of the hemimaxillar surface area occupied by FIL was 6.83%, 4.82%, 2.88%, and 6.52% in rats at age 16, 22, 28, and 34 wk, respectively. Histopathologic changes in FIL varied from localized gingivitis to larger, localized periodontitis-like lesions. These data indicate that FIL are common in rice rats fed standard rodent chow, are slight to mild in severity, and are localized to specific regions in the oral cavity, thus suggesting they may be a suitable model for local maxillary periodontitis when fed standard rodent chow.