LncRNA RP11-400K9.4 Aggravates Cardiomyocytes Apoptosis After Hypoxia/Reperfusion Injury by Targeting miR-423.

Our study was aimed at exploring the roles of lncRNA RP11-400K9.4 (RP11-400K9.4) on hypoxia/reoxygenation (H/R) -induced cardiomyocytes apoptosis. H/R model was constructed in rat primary cardiomyocytes (PC) and H9c2 cells. In this study, the results showed that H/R significantly induced the apoptosis of PC and H9c2 cells. The expression of RP11-400K9.4 was upregulated in H/R-induced PC and H9c2 cells, but miR-423 expression was downregulated. Silencing RP11-400K9.4 could attenuate H/R-induced apoptosis in PC and H9c2 cells. We also found that miR-423 was a potential target of RP11-400K9.4. The effect of silencing RP11-400K9.4 on H/R-induced apoptosis of PC and H9c2 cells was significantly reversed by miR-423 inhibitor transfection. Furthermore, our data confirmed that silencing RP11-400K9.4 promoted the activation of phosphatidylinositol 3-kinase/protein kinase B (PI3K/AKT) and mitogen-activated protein kinase/extracellular signal-regulated kinase (MEK) /extracellular signal-regulated kinase (ERK) pathways and these phenomena can be reversed by miR-423 inhibitor transfection. In conclusion, our study demonstrated that silencing RP11-400K9.4 could alleviate H/R-induced cardiomyocytes damages via suppressing apoptosis by targeting miR-423 with the activation of PI3K/AKT and MEK/ERK signaling pathways.

Protective mechanism of fdft1 in steroid hormone synthesis pathway in SD rats with acute hypoxic injury.

BACKGROUND: The acute hypoxic injury caused by the plain population entering the plateau in a short period of time has become the main cause of endangering the health of the people who rush into the plateau. OBJECTIVE: The study aimed to identify the key genes which participate in resisting the acute hypoxic injury in SD Rats by transcriptomic profile analysis. METHODS: 48 Sprague Dawley (SD) male rats were enrolled and randomly divided into four groups (0h, 24h, 48h, 72h) and housed in hypobaric hypoxia chamber with altitude 6000m for different periods of time to make them acute hypoxic injury. The transcriptomic profile of the lung tissue of the rats was analysed by RNA second-generation sequencing combined with bioinformatics analysis. RESULTS: The results of GO and KEGG function classification analysis revealed that the differential expression genes enriched in steroid hormone synthesis pathway especially in 48h group compared to F0 group. Further analysis revealed that Farnesyl Diphosphate Farnesyl Transferase 1 (fdft1) gene encoding a rate-limiting enzyme in steroid hormone synthesis pathway was significant differently expressed between the groups. The expression levels of fdft1 gene were further verified by RT-PCR and Western-blot methods. CONCLUSIONS: The results suggest that fdft1 gene plays an important role in responding to acute hypoxic injury by regulating steroid hormone biosynthesis.

The deficiency of myelin in the mutant taiep rat induces a differential immune response related to protection from the human parasite Trichinella spiralis.

Taiep rat is a myelin mutant with a progressive motor syndrome characterized by tremor, ataxia, immobility episodes, epilepsy and paralysis of the hindlimbs. Taiep had an initial hypomyelination followed by a progressive demyelination associated with an increased expression of some interleukins and their receptors. The pathology correlated with an increase in nitric oxide activity and lipoperoxidation. In base of the above evidences taiep rat is an appropriate model to study neuroimmune interactions. The aim of this study was to analyze the immune responses in male taiep rats after acute infection with Trichinella spiralis. Our results show that there is an important decrease in the number of intestinal larvae in the taiep rat with respect to Sprague-Dawley control rats. We also found differences in the percentage of innate and adaptive immune cell profile in the mesenteric lymphatic nodes and the spleen that correlated with the demyelination process that took place on taiep subjects. Finally, a clear pro-inflammatory cytokine pattern was seen on infected taiep rats, that could be responsible of the decrement in the number of larvae number. These results sustain the theory that neuroimmune interaction is a fundamental process capable of modulating the immune response, particularly against the parasite Trichinella spiralis in an animal model of progressive demyelination due to tubulinopathy, that could be an important mechanism for the clinical course of autoimmune diseases associated with parasite infection.

Proper mechanical stress promotes femoral head recovery from steroid-induced osteonecrosis in rats through the OPG/RANK/RANKL system.

BACKGROUND: Long-term use of steroid may lead to osteonecrosis of the femoral head (ONFH). Mechanical stress may help bone formation and remodeling. This study aimed to probe the role of mechanical stress in the femoral head recovery in rats. METHODS: Rat models with ONFH were induced by steroid. Rats were subjected to different levels of mechanical stress (weight-bearing training), and then the morphology and bone density of femoral head of rats were measured. The mRNA and protein levels of the OPG/RANK/RANKL axis in rat femoral head were assessed. Gain- and loss-of function experiments of OPG were performed to identify its role in femoral head recovery following stress implement. The ex vivo cells were extracted and the effects of stress and OPG on osteogenesis in vitro were explored. RESULTS: Steroid-induced ONFH rats showed decreased bone density and increased bone spaces, as well as necrotic cell colonies and many cavities in the cortical bones and trabeculars. Proper mechanical stress or upregulation of OPG led to decreased RANK/RANKL expression and promoted femoral head recovery from steroid-induced osteonecrosis. However, excessive mechanical stress might impose too much load on the femurs thus leading even retard femoral head recovery process. In addition, the in vitro experimental results supported that proper stress and overexpression of OPG increased the osteogenesis of ex vivo cells of femoral head. CONCLUSION: This study provided evidence that proper mechanical stress promoted femoral head recovery from steroid-induced osteonecrosis through the OPG/RANK/RANKL system, while overload might inhibit the recovery process. This study may offer novel insights for ONFH treatment.

Efficient derivation of knock-out and knock-in rats using embryos obtained by in vitro fertilization.

Rats are effective model animals and have contributed to the development of human medicine and basic research. However, the application of reproductive engineering techniques to rats is not as advanced compared with mice, and genome editing in rats has not been achieved using embryos obtained by in vitro fertilization (IVF). In this study, we conducted superovulation, IVF, and knock out and knock in using IVF rat embryos. We found that superovulation effectively occurred in the synchronized oestrus cycle and with anti-inhibin antiserum treatment in immature rats, including the Brown Norway rat, which is a very difficult rat strain to superovulate. Next, we collected superovulated oocytes under anaesthesia, and offspring derived from IVF embryos were obtained from all of the rat strains that we examined. When the tyrosinase gene was targeted by electroporation in these embryos, both alleles were disrupted with 100% efficiency. Furthermore, we conducted long DNA fragment knock in using adeno-associated virus and found that the knock-in litter was obtained with high efficiency (33.3-47.4%). Thus, in this study, we developed methods to allow the simple and efficient production of model rats.

MiR-543-3p promotes locomotor function recovery after spinal cord injury by inhibiting the expression of tumor necrosis factor superfamily member 15 in rats.

OBJECTIVE: To explore the effect of miR-543-3p on the recovery of locomotor function after spinal cord injury (SCI) by regulating tumor necrosis factor superfamily member 15 (TNFSF15) mediated inflammation and apoptosis. MATERIALS AND METHODS: Macrophages were isolated from the abdominal cavity of 2-3 months old Sprague-Dawley (SD) rats and cultured. The levels of miR-543-3p, tumor necrosis factor superfamily member 15 (TNFSF15), TNF-like molecule 1A (TL1A) and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) after transfection of miR-92b-5p into activated macrophages were detected by quantitative Real-time polymerase chain reaction (qRT-PCR). Moreover, the mRNA expressions of miR-543-3p, TNFSF15, TL1A and NF-κB after SCI in rats were detected by qRT-PCR. Meanwhile, the protein expressions of tumor necrosis factor (TNF-α), interleukin-1 ß (IL-1ß) and Caspase8 were detected by Western blot. After intrathecal injection of miR-543-3p mimics, the mRNA expressions of miR-543-3p, TNFSF15, TL1A and NF-κB and the protein expressions of TNF-α, IL-1ß and Caspase8 in spinal cord injured area of mice were measured by qRT-PCR and Western blot, respectively. Basso Beattie Bresnahan (BBB) locomotor rating scale was used to determine the recovery of locomotor function after spinal cord injury and injection of miR-543-3p mimics. RESULTS: Compared with inactivated cells, the expression of miR-543-3p in activated macrophages was significantly declined. However, the levels of TNFSF15, TL1A and NF-κB were significantly elevated. The expressions of TNFSF15, TL1A and NF-κB were remarkably downregulated after transfection of miR-543-3p. In addition, the level of miR-543-3p was significantly downregulated, accompanied by an increase in TNFSF15, TL1A and NF-κB in SCI rats. Accordingly, the protein levels of TNF-α and IL-1ß and Caspase8 were also significantly increased. However, the expressions of TNFSF15, TL1A and NF-κB were significantly down-regulated in rats injected with miR-543-3p mimics, whereas the protein levels of TNF-α and IL-1ß and Caspase8 were significantly suppressed. Finally, compared with SCI group, the recovery of locomotor function in miR-543-3p mimics administration group was significantly improved. CONCLUSIONS: After SCI, miR-543-3p can inhibit the activity of NF-κB by suppressing the inflammatory aggravation of TNFSF15 and decreasing its product TL1A. MiR-543-3p leads to the improvement of neuron protection and locomotor function via attenuating inflammatory reaction and cell apoptosis.

Quantitative Analysis of UDP-Glucuronosyltransferase Ugt1a and Ugt2b mRNA Expression in the Rat Liver and Small Intestine: Sex and Strain Differences.

UDP-glucuronosyltransferases (UGTs) catalyze the glucuronidation of numerous endogenous and exogenous compounds to facilitate their excretion from the body. Because rats are commonly used in nonclinical studies, information regarding UGT species differences between rats and humans would be helpful for understanding human pharmacokinetics. In this study, we determined the absolute mRNA expressions of Ugt isoforms in the liver and small intestine of male and female Sprague-Dawley, Fischer 344, and Wistar rats. The sum of the mRNA levels of Ugt isoforms expressed in the liver was significantly (P < 0.005) higher than that in the small intestine regardless of the strain and sex. Ugt2b mRNA levels represented approximately 80% of total Ugt mRNA levels in the liver, whereas Ugt1a mRNA levels accounted for almost 90% in the small intestine. Ugt2b2 mRNA was specifically expressed in Wistar rat liver, resulting in 2-fold higher expression of total hepatic Ugt mRNA in Wistar rats than that in the other strains. Wistar rats showed prominently higher Ugt2b3 and Ugt2b8 mRNA levels in the small intestine than the other strains. The difference between sexes was remarkable with regard to hepatic Ugt1a10 in any of the strains, although slight differences between sexes were also observed in multiple Ugt isoforms. Taken together, this study revealed sex and strain differences in mRNA levels of rat Ugts. The data shown here would be useful for the selection of rat strains in nonclinical studies.

The Effect of Chemerin on Cardiac Parameters and Gene Expressions in Isolated Perfused Rat Heart

Background: Chemerin is a novel chemoattractant adipokine expressed in cardiovascular system, and its receptor has been detected in the epicardial adipose tissue. Aims: To determine the effects of chemerin on the cardiac parameters and gene expressions in the isolated perfused rat heart. Study Design: Animal experiment. Methods: The hearts were retrogradely perfused with Langendorff technique to measure the cardiac parameters. The experimental groups were acutely treated with 10, 100, and 1000 nM doses of chemerin. Another group was given 10 µM L-nitric oxide synthase inhibitor for 5 min before 1000 nM chemerin administration. The real-time polymerase chain reaction was performed for detecting the expression of target genes. Results: All doses of chemerin significantly decreased the left ventricular developed pressure (max 35.33 Δ%, p<0.001), and +dP/dtmax (max 31.3 Δ%, p<0.001), which are the indexes of cardiac contractile force. In addition, 1000 nM chemerin reduced the coronary flow (max 31 Δ%, p<0.001). N(W)-nitro-L-arginine methyl ester antagonized the negative inotropic effect of chemerin on contractility. Chemerin induced a 2.16-fold increase in endothelial nitric oxide synthase mRNA and increased the cyclic guanosine monophosphate levels (p<0.001) but decreased the PI3Kγ gene expression (1.8-fold, p<0.001). Furthermore, all doses of chemerin decreased the CaV1.2 gene expression (1.69-fold, p<0.001). Conclusion: Acute chemerin treatment induces a negative inotropic action with the involvement of nitric oxide pathway, CaV1.2, and PI3Kγ on isolated rat heart.

Screening and Function Analysis of MicroRNAs Involved in Exercise Preconditioning-Attenuating Pathological Cardiac Hypertrophy.

Exercise preconditioning (EP) attenuates pathological cardiac hypertrophy by increasing the functional capacity of the cardiovascular system; however, the underlying molecular mechanisms remain unclear. MicroRNAs (miRNAs) play important roles in various physiological and pathological processes by regulating the expression of the targeted gene. In this study, we aimed to screen the miRNAs involved in EP-attenuating pathological cardiac hypertrophy. The histological and echocardiographic parameters assessment showed that pathological cardiac hypertrophy induced by transverse aortic constriction (TAC) was significantly alleviated in EP treated rats. The left ventricular tissues (n = 3) from Sham, TAC and EP + TAC groups were subjected to small RNA deep sequencing. A total of 570 known mature miRNAs and 530 putative novel miRNAs were detected. DEGseq analysis showed that there were 37 and 88 differentially expressed miRNAs in the comparisons of TAC versus Sham and EP + TAC versus TAC, respectively. Among them, EP treatment could relieve the expression changes of 32 miRNAs, which were supposed to be involved in EP-attenuating pathological cardiac hypertrophy. After miRNAs target genes prediction by miRDB algorithm, pathway analysis showed that the most frequently represented pathways were involved in Calcium signaling pathway and MAPK signaling pathway. The results would provide valuable clues to finding therapeutic targets for the treatment of pathological cardiac hypertrophy.

Biliary tract external drainage improves inflammatory mediators and pathomorphology of the intestine, liver, and lung in septic rats.

BACKGROUND: To investigate the effect of biliary tract external drainage (BTED) on inflammatory mediators and pathomorphism of intestine, liver, and lung in septic rats. METHOD: 48 SD rats (n = 8 per group) were randomized into six groups: control, sepsis, sepsis plus BTED, normal bile (obtained from eight healthy rats), and septic bile infusion for 6 hours respectively to test the effects of BTED bile infusion on cytokines' expression and tissue injury in the intestine, liver, and lung of septic/normal rats. Co-cultivation of intestinal epithelial cells (IEC-6) with bile for 12 hours was performed to evaluate the potential cytotoxicity of septic bile. Survival rate for sepsis plus BTED rats was detected compared with sepsis without BTED group (n = 20 per group) at 24, 48, and 72 hours, respectively. RESULTS: BTED for 6 hours significantly reduced the mRNA expression levels of tumor necrosis factor alpha (TNF-α) and IL-1ß (all p < 0.05 vs. sepsis group), whereas mRNA expression of TNF-α and IL-1ß in the intestine was increased after 6 hours' septic bile infusion compared with normal bile infusion group (all p < 0.05). TNF-α concentration in septic bile was significantly higher than that in the control group (p < 0.001). Tissue injury was significantly attenuated after 6 hours' BTED. CONCLUSIONS: BTED can significantly restrain the mRNA expression of TNF-α and IL-1ß in the intestine, liver, and lung and attenuate histological damage in septic rats.

Methylation pattern variation between goats and rats during the onset of puberty.

Puberty is initiated by increased pulsatile gonadotropin-releasing hormone (GnRH) release from the hypothalamus. Epigenetic repression is thought to play a crucial role in the initiation of puberty, although the existence of analogous changes in methylation patterns across species is unclear. We analysed mRNA expression of DNA methyltransferases (DNMTs) and methyl-binding proteins (MBPs) in goats and rats by quantitative real-time PCR (qRT-PCR). DNA methylation profiles of hypothalamic were determined at the pre-pubertal and pubertal stages by bisulphite sequencing. In this study, expression of DNMTs and MBPs mRNA showed different patterns in goats and rats. Global methylation variation was low in goats and rats, and the profile remained stable during puberty. Gene ontology (GO) and Kyoto Encyclopedia of Gene and Genomes (KEGG) pathway analysis revealed the involvement of 62 pathways in puberty in goats and rats including reproduction, type I diabetes mellitus and GnRH signalling pathways and found that Edn3, PTPRN2 and GRID1 showed different methylation patterns during puberty in goats and rats and similar variation patterns for Edn3 and PTPRN2 were showed. These indicated that Edn3 and PTPRN2 would play a role in the timing of puberty. This study provides evidence of the epigenetic control of puberty.

Isoflurane is a suitable alternative to ether for anesthetizing rats prior to euthanasia for gene expression analysis.

Diethyl ether (ether) had been widely used in Japan for anesthesia, despite its explosive properties and toxicity to both humans and animals. We also had used ether as an anesthetic for euthanizing rats for research in the Toxicogenomics Project (TGP). Because the use of ether for these purposes will likely cease, it is required to select an alternative anesthetic which is validated for consistency with existing TGP data acquired under ether anesthesia. We therefore compared two alternative anesthetic candidates, isoflurane and pentobarbital, with ether in terms of hematological findings, serum biochemical parameters, and gene expressions. As a result, few differences among the three agents were observed. In hematological and serum biochemistry analysis, no significant changes were found. In gene expression analysis, four known genes were extracted as differentially expressed genes in the liver of rats anesthetized with ether, isoflurane, or pentobarbital. However, no significant relationships were detected using gene ontology, pathway, or gene enrichment analyses by DAVID and TargetMine. Surprisingly, although it was expected that the lung would be affected by administration via inhalation, only one differentially expressed gene was extracted in the lung. Taken together, our data indicate that there are no significant differences among ether, isoflurane, and pentobarbital with respect to effects on hematological parameters, serum biochemistry parameters, and gene expression. Based on its smallest affect to existing data and its safety profile for humans and animals, we suggest isoflurane as a suitable alternative anesthetic for use in rat euthanasia in toxicogenomics analysis.

Effect of genetic strain and gender on age-related changes in body composition of the laboratory rat.

Body fat serves as a storage compartment for lipophilic pollutants and affects the pharmacokinetics of many toxic chemicals. Understanding how body fat varies with gender, strain, and age may be essential for development of experimental models to study mechanisms of toxicity. Nuclear magnetic resonance (NMR)-based analysis serves as a noninvasive means of assessing proportions of fat, lean, and fluid in rodents over their lifetime. The aim of this study was to track changes in body composition of male and female Long-Evans (LE), Sprague-Dawley (SD), Fischer (F334), and Brown Norway (BN) rats from postweaning over a >2-yr period. Percent fat of preweaned LE and SD rats was markedly higher compared to the other strains. LE and SD strains displayed marked increases in body fat from weaning to 8 mo of age. Postweaned F344 male and females showed relatively low levels of percent fat; however, at 2 yr of age percent fat of females was equal to that of SD and LE in females. BN rats showed the highest levels of lean tissue and lowest levels of fat. Percent fat of the BN strain rose at the slowest rate as they aged. Percent fluid was consistently higher in males for all strains. Females tended to have higher percent fat than males in LE, SD, and F344 strains. Assessing changes in body fat as well as lean and fluid of various strains of male and female rats over their lifetime may prove useful in many research endeavors, including pharmacokinetics of lipophilic toxicants, mechanisms underlying obesity, and metabolic disorders.

Strain differences in cytochrome P450 mRNA and protein expression, and enzymatic activity among Sprague Dawley, Wistar, Brown Norway and Dark Agouti rats.

Rat cytochrome P450 (CYP) exhibits inter-strain differences, but their analysis has been scattered across studies under different conditions. To identify these strain differences in CYP more comprehensively, mRNA expression, protein expression and metabolic activity among Wistar (WI), Sprague Dawley (SD), Dark Agouti (DA) and Brown Norway (BN) rats were compared. The mRNA level and enzymatic activity of CYP1A1 were highest in SD rats. The rank order of Cyp3a2 mRNA expression mirrored its protein expression, i.e., DA>BN>SD>WI, and was similar to the CYP3A2-dependent warfarin metabolic activity, i.e., DA>SD>BN>WI. These results suggest that the strain differences in CYP3A2 enzymatic activity are caused by differences in mRNA expression. Cyp2b1 mRNA levels, which were higher in DA rats, did not correlate with its protein expression or enzymatic activity. This suggests that the strain differences in enzymatic activity are not related to Cyp2b1 mRNA expression. In conclusion, WI rats tended to have the lowest CYP1A1, 2B1 and 3A2 mRNA expression, protein expression and enzymatic activity among the strains. In addition, SD rats had the highest CYP1A1 mRNA expression and activity, while DA rats had higher CYP2B1 and CYP3A2 mRNA and protein expression. These inter-strain differences in CYP could influence pharmacokinetic considerations in preclinical toxicological studies.

Complete sequence of an ovarian cancer inbred Sprague-Dawley rat model mitochondrial genome.

The Sprague-Dawley rat strain is a commonly used model for ovarian cancer disease study. We sequenced this rat strain mitochondrial genome for the first time (GenBank Accession No. KM114604). Its mitogenome was 16,312 bp and coding 13 protein-coding genes, 2 ribosomal RNA genes and 22 transfer RNA genes. A total of 96 SNPs were examined when compared to reference BN sequence.

Complete mitochondrial genome sequence and mutations of the hepatocellular carcinoma model inbred Sprague-Dawley strain.

In the present work we undertook the complete mitochondrial genome sequencing of an important hepatocellular carcinoma model inbred Sprague-Dawley strain for the first time. The total length of the mitogenome was 16,308 bp. It harbored 13 protein-coding genes, two ribosomal RNA genes, 22 transfer RNA genes and one non-coding control region (D-loop region). The mutation events were also reported.

Borna disease virus infection impacts microRNAs associated with nervous system development, cell differentiation, proliferation and apoptosis in the hippocampi of neonatal rats.

MicroRNAs (miRNAs) regulate gene expression by inhibiting transcription or translation and are involved in diverse biological processes, including development, cellular differentiation and tumor generation. miRNA microarray technology is a high­throughput global analysis tool for miRNA expression profiling. Here, the hippocampi of four borna disease virus (BDV)­infected and four non­infected control neonatal rats were selected for miRNA microarray and bioinformatic analysis. Reverse transcription quantitative polymerase chain reaction (RT­qPCR) analysis was subsequently performed to validate the dysregulated miRNAs. Seven miRNAs (miR­145*, miR­146a*, miR­192*, miR­200b, miR­223*, miR­449a and miR­505), showed increased expression, whereas two miRNAs (miR­126 and miR­374) showed decreased expression in the BDV­infected group. By RT­qPCR validation, five miRNAs (miR­126, miR­200b, miR­374, miR­449a and miR­505) showed significantly decreased expression (P<0.05) in response to BDV infection. Biocarta pathway analysis predicted target genes associated with 'RNA', 'IGF1mTOR', 'EIF2', 'VEGF', 'EIF', 'NTHI', 'extrinsic', 'RB', 'IL1R' and 'IGF1' pathways. Gene Ontology analysis predicted target genes associated with 'peripheral nervous system development', 'regulation of small GTPase-mediated signal transduction', 'regulation of Ras protein signal transduction', 'aerobic respiration', 'membrane fusion', 'positive regulation of cell cycle', 'cellular respiration', 'heterocycle metabolic process', 'protein tetramerization' and 'regulation of Rho protein signal transduction' processes. Among the five dysregulated miRNAs identified by RT­qPCR, miR­126, miR­200b and miR­449a showed a strong association with nervous system development, cell differentiation, proliferation and apoptosis.

Polycystic kidney disease in Sprague-Dawley rats.

Polycystic kidney disease (PKD) is a cystic genetic disorder of the kidneys which is typically associated with cystic bile duct dilatation in the liver in humans, and domestic and laboratory animals. In humans, there are two types of PKD, autosomal dominant polycystic kidney disease (ADPKD) and autosomal recessive polycystic kidney disease (ARPKD). ADPKD is caused by mutations in PKD1 or PKD2 gene while ARPKD is caused by mutation or loss of the PKHD1 (polycystic kidney and hepatic disease 1) gene. Here we report a morphologically confirmed case of spontaneous PKD in a Sprague-Dawley rat in which anatomic pathology examination revealed numerous cystic changes in the kidney and liver. Lesions consisted of marked cystic dilatations of renal tubules, and moderate cystic dilatations of intrahepatic bile ducts with portal fibrosis. We present detailed histologic features of the spontaneous PKD and compare them with disease model rats carrying an autosomal recessive PKHD 1 gene mutation.

Chondrocyte density, proteoglycan content and gene expressions from native cartilage are species specific and not dependent on cartilage thickness: a comparative analysis between rat, rabbit and goat.

BACKGROUND: In many pre-clinical studies of cartilage tissue, it has been generally assumed that the major difference of the tissue between the species is the tissue thickness, which is related to the size of the animal itself. At present, there appear to be lack of studies demonstrating the relationship between chondrocyte densities, protein content, gene expressions and cartilage thickness in the various animal models that are commonly used. The present study was conducted to determine whether or not chondrocyte density, proteoglycan/protein content and selective chondrocyte gene expression are merely related to the cartilage thickness (thus animal size), and not the intrinsic nature of the species being investigated. Mature animals (rabbit, rats and goats) were sacrificed for their hind knee cartilages. Image analyses were performed on five consecutive histological sections, sampled from three pre-defined locations at the lateral and medial femoral condyles. Cartilage thickness, chondrocyte density, Glycosaminoglycan (GAGs)/protein content and gene expression levels for collagen II and SOX-9 were compared across the groups. Correlation analysis was done between cartilage thickness and the other variables. RESULTS: The mean cartilage thickness of rats, rabbits and goats were 166.5 ± 10.9, 356.2 ± 25.0 907.5 ± 114.6 µm, respectively. The mean cartilage cell densities were 3.3 ± 0.4×10(-3) for rats, 2.6 ± 0.3×10(-3) for rabbits and 1.3 ± 0.2×10(-3) cells/µm2 for goats. The mean µg GAG/mg protein content were 23.8 ± 8.6 in rats, 20.5 ± 5.3 in rabbits and 328.7 ± 64.5 in goats; collagen II gene expressions were increased by 0.5 ± 0.1 folds in rats; 0.6 ± 0.1 folds in rabbits, and 0.1 ± 0.1 folds in goats, whilst the fold increase of SOX-9 gene expression was 0.5 ± 0.1 in rats, 0.7 ± 0.1 in rabbits and 0.1 ± 0.0 in goats. Cartilage thickness correlated positively with animals' weight (R2 =0.9856, p = 0.001) and GAG/protein content (R2 =0.6163, p = <0.001). Whereas, it correlates negatively with cell density (R2 = 0.7981, p < 0.001) and cartilage gene expression levels (R2 = 0.6395, p < 0.001). CONCLUSION: There are differences in the composition of the articular cartilage in diverse species, which are not directly dependent on the cartilage thickness of these animals but rather the unique characteristics of that species. Therefore, the species-specific nature of the cartilage tissue should be considered during any data interpretation.

The expression patterns of Reg IV gene in normal rat reproduction system.

Reg IV, the latest member of the regenerating gene family, has been documented in different tissues of human and rat, such as the colon, small intestine, stomach, and pancreas. Expression of Reg IV gene in distinct cell types has been correlated with its various functions in regeneration, cell growth and survival, proliferation and differentiation, cell adhesion, and resistance to apoptosis. However, there was no evidence to show whether the Reg IV protein is present in the reproductive system of normal rat. The aim of this study was to reveal the expression patterns of Reg IV in rat ovary and uterus. The expression of Reg IV was analyzed by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot at mRNA and protein levels, respectively. The localization of Reg IV protein within rat ovary and uterus was investigated by immunohistochemistry (IHC). Our results showed that the expression of Reg IV in ovary was significantly higher than that in the uterus. The strong immunoreactive signals of Reg IV was observed in granulosa cells and oocytes of ovarian follicles, corpus luteum, and interstitial cells in rat ovary; only weak signals were detected in luminal and gland epithelium of rat endometrium. These findings first demonstrate the expression of Reg IV in ovary and uterus of the healthy rat at both mRNA and protein levels. It provides an evidence of Reg IV expression in rat reproductive system, which may help elucidate a potential role in cell growth and proliferation of reproductive system.