Kanchana Kengkoom

High Dietary Cholesterol Masks Type 2 Diabetes-Induced Osteopenia and Changes in Bone Microstructure in Rats

Type 2 diabetes mellitus (T2DM) often occurs concurrently with high blood cholesterol or dyslipidemia. Although T2DM has been hypothesized to impair bone microstructure, several investigations showed that, when compared to age-matched healthy individuals, T2DM patients had normal or relatively high bone mineral density (BMD). Since cholesterol and lipids profoundly affect the function of osteoblasts and osteoclasts, it might be cholesterol that obscured the changes in BMD and bone microstructure in T2DM. The present study, therefore, aimed to determine bone elongation, epiphyseal histology, and bone microstructure in non-obese T2DM Goto-Kakizaki rats treated with normal (GK-ND) and high cholesterol diet. We found that volumetric BMD was lower in GK-ND rats than the age-matched wild-type controls. In histomorphometric study of tibial metaphysis, T2DM evidently suppressed osteoblast function as indicated by decreases in osteoblast surface, mineral apposition rate, and bone formation rate in GK-ND rats. Meanwhile, the osteoclast surface and eroded surface were increased in GK-ND rats, thus suggesting an activation of bone resorption. T2DM also impaired bone elongation, presumably by retaining the chondrogenic precursor cells in the epiphyseal resting zone. Interestingly, several bone changes in GK rats (e.g., increased osteoclast surface) disappeared after high cholesterol treatment as compared to wild-type rats fed high cholesterol diet. In conclusion, high cholesterol diet was capable of masking the T2DM-induced osteopenia and changes in several histomorphometric parameters that indicated bone microstructural defect. Cholesterol thus explained, in part, why a decrease in BMD was not observed in T2DM, and hence delayed diagnosis of the T2DM-associated bone disease.

Pulmonary Edema Due to Oral Gavage in a Toxicological Study Related to Aquaporin-1, -4 and -5 Expression

A one-time oral gavage can be enough to cause of alveologenic edema with higher expression of AQP-1 and -4 than that with repeated-dose oral gavage, which caused both profound perivascular edema and hydrostatic pressure edema, while AQP-5 was similarly expressed. The alteration of AQPs expression was probably related to alveolar fluid clearance across the alveolar and bronchiolar epithelium in different stages of lung injury. The results clarified the type of lung edema in acute and sub-chronic toxicity studies without treatment related effect of tested material. The pathogenesis of pulmonary edema due to oral gavage toxicological study is associated with the cellular immune response to the reflux materials. Mast cell and leukocyte accumulation may contribute to increase vascular permeability leading to permeability edema. The increase in alveolar septum epithelium, perivascular and peribronchial cuffing, accumulation alveolar lipid containing macrophage and medial hyperplasia of the pulmonary artery might have been caused to increase airway resistance, which resulted in hydrostatic pressure edema.

Prevalence of Helicobacter in Laboratory Micein Thailand

Prevalence of Helicobacter is mostly unknown in laboratory animals in Thailand. The 221 mice feces/cecum from 8 universities, 2 pharmaceutical companies and 3 research institutions in Thailand were surveyed for the prevalence and distribution of Helicobacter species by using the Electrochemical DNA chip. Helicobacter were detected 23/46 samples in Specific Pathogen Free (SPF) and 168/175 in conventional condition. Prevalence of Helicobacter were 98%, 96%, 92% and 78% in South (n=40), Northeast (n=40), North (n=25) and Central area (n=116), respectively. Only Central area holds SPF facility resulting in Helicobacter prevalence that seems to be lower than other areas. Three species of Helicobacter were detected in feces/cecum samples by sequence analysis: H. rodentium (67.0%, 148 samples), Helicobacter sp. MIT 01-6451 (15.4%, 34 samples), and unidentified Helicobacter species (14.1%, 9 samples). The results suggested that H. rodentium is the most common species of Helicobacter in laboratory mice in Thailand.

Defective Bone Microstructure in Hydronephrotic Mice: A Histomorphometric Study in ICR/Mlac‐hydro Mice

Chronic renal impairment can lead to bone deterioration and abnormal bone morphology, but whether hydronephrosis is associated with bone loss remains unclear. Herein, we aimed to use computer‐assisted bone histomorphometric technique to investigate microstructural bone changes in Imprinting Control Region (ICR) mice with a spontaneous mutation that was associated with bilateral nonobstructive hydronephrosis (ICR/Mlac‐hydro). The results showed that 8‐week‐old ICR/Mlac‐hydro mice manifested decreases in trabecular bone number and thickness, and an increased trabecular separation, thereby leading to a reduction in trabecular bone volume compared with the wild‐type mice. Furthermore, histomorphometric parameters related to both bone resorption and formation, that is, eroded surface, osteoclast surface, and osteoblast surface, were much lower in ICR/Mlac‐hydro mice than in the wild type. A decrease in moment of inertia was found in ICR/Mlac‐hydro mice, indicating a decrease in bone strength. In conclusion, ICR/Mlac‐hydro mice exhibited trabecular bone loss, presumably caused by marked decreases in both osteoblast and osteoclast activities, which together reflected abnormally low bone turnover. Thus, this mouse strain appeared to be a valuable model for studying the hydronephrosis‐associated bone disease. Anat Rec, 297:208–214, 2014.

Expression of aquaporin-1, -2 and -4 in mice with a spontaneous mutation leading to hydronephrosis.

This study investigates the expression of aquaporin-1, -2 and -4 in mice with a spontaneously-arising mutation that leads to hydronephrosis (ICR/Mlac-hydro mice). The mutant mice developed bilateral non-obstructive hydronephrosis without evidence of interstitial fibrosis or glomerulosclerosis. The mice had no abnormality in blood urea nitrogen or creatinine concentrations or in urine specific gravity. Despite the severity of the renal damage the mice grew and reproduced normally. Kidneys from the mutant mice had reduced expression of all three aquaporins compared with wild type mice. The reduction in aquaporin was proportional to the degree of hydronephrosis, but this change did not appear to be associated with disturbance of urinary function.

Chronic Ingestion of High Dosed Phikud Navakot Extraction Induces Mesangiolysis in Rats with Alteration of AQP1 and Hsp60 Expressions

Phikud Navakot (PN) is commonly used in Thai traditional medicine for alleviation of cardiovascular and cerebrovascular symptoms; however little is known about the chronic toxicity effects of the extracts from the herbs in PN. Repeated extraction doses of 10, 100, and 1,000 mg/kg/day were randomly administered to both male and female Sprague Dawley rats for 12 months. Histopathological study revealed that mesangiolysis was predominately found at the highest dose. Aquaporin 1 (AQP1) expression in the mesangiolytic glomeruli was significantly lower than in the intact glomeruli. This may be relevant to an imbalance of vascular function manifested by AQP1 alteration. In the mesangiolytic glomeruli, 60 kDa heat shock protein (Hsp60) was significantly upregulated on the endothelial lining cells of aneurysm and vascular cyst. Hsp60 increase may be related to endothelial cell damage due to its intracellular protective role. Blood urea nitrogen and creatinine levels remained within their normal range indicating well-functioning renal reserve function. In conclusion, high dosed PN may affect the endothelium leading to inability of vascular permeability and consequence to mesangiolysis. Our results suggest that only a high dose of chronic oral administration of PN is relatively toxic in association with mesangiolysis. The NOAEL was determined to be 100 mg/kg/day.

Insulin does not rescue cortical and trabecular bone loss in type 2 diabetic Goto-Kakizaki rats

In type 2 diabetes mellitus (T2DM), the decreased bone strength is often associated with hyperglycemia and bone cell insulin resistance. Since T2DM is increasingly reported in young adults, it is not known whether the effect of T2DM on bone would be different in young adolescents and aging adults. Here, we found shorter femoral and tibial lengths in 7-month, but not 13-month, Goto-Kakizaki (GK) T2DM rats as compared to wild-type rats. Bone µCT analysis showed long-lasting impairment of both cortical and trabecular bones in GK rats. Although insulin treatment effectively improved hyperglycemia, it was not able to rescue trabecular BMD and cortical thickness in young adult GK rats. In conclusion, insulin treatment and alleviation of hyperglycemia did not increase BMD of osteopenic GK rats. It is likely that early prevention of insulin resistance should prevail over treatment of full-blown T2DM-related osteopathy.

In Vitro Protective Effect of Phikud Navakot Extraction on Erythrocyte.

Phikud Navakot (PN), Thai herbal remedy in National List of Essential Medicines, has been claimed to reduce many cardiovascular symptoms especially dizziness and fainting. Apart from blood supply, erythrocyte morphology, in both shape and size, is one of the main consideration factors in cardiovascular diseases and may be affected by vascular oxidative stress. However, little is known about antioxidative property of PN on erythrocyte to preserve red blood cell integrity. In this study, 1,000 μM hydrogen peroxide-induced oxidative stress was conducted on sheep erythrocyte. Three doses of PN (1, 0.5, and 0.25 mg/mL) and 10 μM of ascorbic acid were compared. The released hemoglobin absorbance was measured to demonstrate hemolysis. Electron microscopic and immunohistochemical studies were also performed to characterize dysmorphic erythrocyte and osmotic ability in relation to aquaporin- (AQP-) 1 expression, respectively. The results revealed that all doses of PN and ascorbic acid decreased the severity of dysmorphic erythrocyte, particularly echinocyte, acanthocyte, knizocyte, codocyte, clumping, and other malformations. However, the most effective was 0.5 mg/mL PN dosage. In addition, hydrostatic pressure may be increased in dysmorphic erythrocyte in association with AQP-1 upregulation. Our results demonstrated that PN composes antioxidative effect to maintain the integrity and osmotic ability on sheep erythrocyte.

Omeprazole preserves the RER in chief cells and enhances re‑epithelialization of parietal cells with SOD and AQP‑4 up‑regulation in ethanol‑induced gastritis rats

Gastric mucosal cells, particularly parietal and chief cells, are usually affected by exogenous, and endogenous stimuli-induced gastritis. The integrity of these cells and their alterations are involved in the pathogenesis of numerous gastric disorders. Omeprazole, a gastric acid secretion blocker, is commonly used for gastrointestinal diseases due to its antioxidative stress and anti-inflammatory properties. Little is known regarding how omeprazole modulates the re-epithelialized effect on gastric mucosal cells associated with gastrointestinal disorders. The present study aimed to determine whether omeprazole attenuates parietal and chief cell damage in association with its antioxidative property. An in vivo ethanol-induced gastritis rat model was used. Histopathological, scanning and transmission electron microscopic, and immunohistochemical studies were performed. The results revealed that omeprazole improved the gastric mucosal surface, and reduced the severity of mucosal inflammation and hemorrhaging. Notably, ethanol-induced gastritis caused dysmorphic rough endoplasmic reticulum (RER) in chief cells, which was accompanied by mitochondrial swelling. This alteration was modulated by omeprazole due to its antioxidant effect characterized by upregulation of superoxide dismutase in gastric mucosal cells. In addition, expression of aquaporin-4 was increased in the omeprazole treatment group, which may be due to the expansion of regenerative parietal cells and acid suppression. The results of the present study suggest that omeprazole preserves the RER in chief cells and enhances parietal cell regeneration through its antioxidative property by exerting anti-inflammatory effects.