Kidney injury molecule-1 and urinary gamma-glutamyl transferase as biomarkers of acute kidney injury in cats.

OBJECTIVES: To evaluate the concentration of kidney injury molecule-1 and activity of urinary gamma-glutamyl transferase in cats with urethral obstruction and healthy cats. MATERIALS AND METHODS: Blood and urine samples were collected from a group of 15 healthy cats (control group) and a group of 20 cats with urethral obstruction at presentation, and 24 hours and 7 days after unblocking the obstruction. The serum creatinine, urinary creatinine and urinary gamma-glutamyl transferase were measured by spectrophotometry and kidney injury molecule-1 by the sandwich enzyme-linked immunosorbent assay. RESULTS: On presentation, cats with obstruction had serum creatinine concentration and urinary gamma-glutamyl transferase index higher than healthy cats (mean difference 544 µmol/L, 95% confidence intervals 222 to 865 µmol/L, and 0.0022 U/µmol-uCre, 0.00043 to 0.0039 U/µmol-uCre, respectively), urine creatinine concentration lower (mean difference 25,624 µmol/L, 17,329 to 33,919 µmol/L), and no significant difference in the kidney injury molecule-1/urinary creatinine ratio (mean difference 13 pg/µmol-uCre, -33 to 59 pg/µmol-uCre). In the group of cats with urinary obstruction, over time serum creatinine decreased, urine creatinine increased, urinary gamma-glutamyl transferase index did not change significantly, and kidney injury molecule-1/urinary creatinine ratio increased. CLINICAL SIGNIFICANCE: Cats with post-renal obstruction and potential intrinsic renal damage had higher urinary gamma-glutamyl transferase index than healthy cats at the time of presentation and showed increase in kidney injury molecule-1/urinary creatinine ratio over time.

α,ß-Amyrin prevents steatosis and insulin resistance in a high-fat diet-induced mouse model of NAFLD via the AMPK-mTORC1-SREBP1 signaling mechanism.

Nonalcoholic fatty liver disease (NAFLD), characterized by hepatosteatosis and steatohepatitis, is intrinsically related to obesity. Our previous study reported on the anti-obese activity of α,ß-amyrin (AMY), a pentacyclic triterpene isolated from Protium heptaphyllum. This study investigated its ability to prevent fatty liver and the underlying mechanism using the mouse model of NAFLD. NAFLD was induced in male Swiss mice fed a high fat diet (HFD) for 15 weeks. The controls were fed a normal chow diet (ND). The mice were simultaneously treated with AMY at 10 and 20 mg/kg or fenofibrate at 50 mg/kg. Lipid levels along with metabolic and inflammatory parameters were assessed in liver and serum. The liver sections were histologically examined using H&E staining. RT-qPCR and western blotting assays were performed to analyze signaling mechanisms. Mice fed HFD developed severe hepatic steatosis with elevated triglycerides and lipid droplets compared with ND controls. This was associated with a decrease in AMP-activated protein kinase (AMPK) activity, an increase of mechanistic target of rapamycin complex 1 (mTORC1) signaling, and enhanced sterol regulatory element binding protein 1 (SREBP1) expression, which have roles in lipogenesis, inhibition of lipolysis, and inflammatory response. AMY treatment reversed these signaling activities and decreased the severity of hepatic steatosis and inflammatory response, evidenced by serum and liver parameters as well as histological findings. AMY-induced reduction in hepatic steatosis seemed to involve AMPK-mTORC1-SREBP1 signaling pathways, which supported its beneficial role in the prevention and treatment of NAFLD.

α, ß-Amyrin prevents steatosis and insulin resistance in a high-fat diet-induced mouse model of NAFLD via the AMPK-mTORC1-SREBP1 signaling mechanism

Nonalcoholic fatty liver disease (NAFLD), characterized by hepatosteatosis and steatohepatitis, is intrinsically related to obesity. Our previous study reported on the anti-obese activity of α,β-amyrin (AMY), a pentacyclic triterpene isolated from Protium heptaphyllum. This study investigated its ability to prevent fatty liver and the underlying mechanism using the mouse model of NAFLD. NAFLD was induced in male Swiss mice fed a high fat diet (HFD) for 15 weeks. The controls were fed a normal chow diet (ND). The mice were simultaneously treated with AMY at 10 and 20 mg/kg or fenofibrate at 50 mg/kg. Lipid levels along with metabolic and inflammatory parameters were assessed in liver and serum. The liver sections were histologically examined using H&E staining. RT-qPCR and western blotting assays were performed to analyze signaling mechanisms. Mice fed HFD developed severe hepatic steatosis with elevated triglycerides and lipid droplets compared with ND controls. This was associated with a decrease in AMP-activated protein kinase (AMPK) activity, an increase of mechanistic target of rapamycin complex 1 (mTORC1) signaling, and enhanced sterol regulatory element binding protein 1 (SREBP1) expression, which have roles in lipogenesis, inhibition of lipolysis, and inflammatory response. AMY treatment reversed these signaling activities and decreased the severity of hepatic steatosis and inflammatory response, evidenced by serum and liver parameters as well as histological findings. AMY-induced reduction in hepatic steatosis seemed to involve AMPK-mTORC1-SREBP1 signaling pathways, which supported its beneficial role in the prevention and treatment of NAFLD.

Morphological characterization of ckd in cats: Insights of fibrogenesis to be recognized.

Renal fibrosis is characterized by glomerulosclerosis and tubulointerstitial fibrosis and its pathogenesis is associated with the activity of mesenchymal cells (fibroblasts), being essentially characterized by a process of excessive accumulation resulting from the deposition of extracellular matrix components. The aim of this study was to characterize the morphological presentation of chronic and fibrotic lesions in the glomerular, tubular, interstitial, and vascular compartments in feline CKD, as well as the possible participation of myofibroblasts in renal fibrotic processes in this species. Cat kidneys were collected and processed according to the conventional techniques for light microscopy, circular polarization, immunohistochemistry, and electron microscopy. Fibrotic alterations were present in all compartments analyzed. The main findings in the glomerular compartment were different degrees of glomerular sclerosis, synechia formation, Bowman's capsule calcification, in addition to glomerular basement membrane thickening and pericapsular fibrosis. The tubulointerstitial compartment had intense tubular degeneration and the immunostaining in tubular cells for mesenchymal cell markers demonstrated the possibility of mesenchymal epithelial transition and consequent involvement of myofibroblasts in the development of interstitial tubule damage. Infiltration of inflammatory cells, added to vessel thickening and fibrosis, demonstrated the severity and role of inflammation in the development and perpetuation of damage. Thus, we may conclude that fibrotic lesions play a relevant role in feline CKD and the mechanism of perpetuation of these lesions need further elucidation regarding the origin and participation of myofibroblasts and consequent mesenchymal epithelial transition in this species.

Síndrome de Eisenmenger em um gato / Eisenmenger syndrome in cat

A 6-month-old female, 1.0kg, uncastrated female Persian cat was brought to the Veterinary Hospital at State University of Ceara, with a history of dyspnea, prostration, hyporexia and progressive weight loss for a month. On physical examination, systolic cardiac murmur, cyanosis and dyspnea were detected. Unfortunately, the cat died during oxygen therapy. Necropsy examination revealed an increase in cardiac silhouette and ventricular septal defect of 2cm in diameter. Macroscopically the lungs were collapsed, with absent and diffusely reddish blackish crepitus, and the liver with blackish red coalescent multifocal areas, interspersed with lighter areas and lobular pattern with irregular brownish multifocal areas intercepted by brownish areas. Thus, the necropsy results together with the history and physical examination of the animal confirmed the diagnosis of Eisenmenger Syndrome, becoming the report of the first case, in a cat, in Brazil.(AU)

Evaluation of the toxicity and hypoglycemic effect of the aqueous extracts of Cnidoscolus quercifolius Pohl.

Diabetes mellitus is one of the most common chronic degenerative diseases, and it is estimated to increase worldwide to around 415 million and to impact 642 million in 2040. Research shows that some plants are sources of bioactive compounds against diabetes. Thus, the objective of this work was to evaluate the oral toxicity and the hypoglycemic effect of the aqueous extract of the leaves of Cnidoscolus quercifolius Pohl. Diabetes was induced in Swiss mice with streptozotocin and the mice were treated with an aqueous extract of C. quercifolius leaves for a period of 30 days. Phytochemical analysis showed that the extract was rich in flavonoids, catechins and triterpenoid, which did not show any mortality and behavioral alterations in mice treated with 200, 1000, and 2000 mg/kg body weight of the extract for 14 days. Histopathological analysis of organs (kidney, pancreas, liver) from mice treated with the 2000 mg/kg extract revealed no architectural change. In the present study, we found a 29% reduction in glucose levels in animals receiving 200 mg/kg body weight. These results are very promising because they showed that C. quercifolius had a hypoglycemic effect and did not present oral toxicity, thus being a new source of compounds for the control of diabetes.

Serum testosterone, sperm quality, cytological, physicochemical and biochemical characteristics of the prostatic fraction of dogs with prostatomegaly.

Prostatomegaly is a common finding in older non-neutered dogs. This study compared the serum testosterone, sperm quality and characteristics of the prostatic fraction between healthy dogs and dogs with prostatomegaly. Blood samples of ten dogs (five dogs from each group) were taken for serum testosterone measurement. Sperm motility, vigour, concentration, viability, membrane functionality and morphology were analysed in sperm-rich fraction. Osmolality, pH, cell types, and albumin, haemoglobin, acid phosphatase, alkaline phosphatase, glucose, triglycerides, cholesterol, calcium, phosphorus, magnesium and chloride were analysed in prostatic fraction. Dogs with prostatomegaly have the lowest sperm motility, vigour, concentration and functional membrane. Dogs with prostatomegaly have the highest glucose, triglycerides and cholesterol. Glucose was the only constituent positively correlated with serum testosterone and prostate volume. It can be concluded that dogs with prostatomegaly have poorer sperm quality, and glucose, triglycerides and cholesterol in prostatic fraction can be used as prostatomegaly biomarkers.

Polarization microscopy as a tool for quantitative evaluation of collagen using picrosirius red in different stages of CKD in cats.

Chronic kidney disease (CKD) is a relevant disease in feline clinic. The tubulointerstitial damage, with collagen deposition and fibrosis, is an important result of this process. The aim of this study was to quantify and correlate the deposition of collagen and severity of interstitial fibrosis (IF) in the kidney from cats in different stages of CKD. Kidney fragments from 10 adult cats with CKD were analyzed and stained by Masson's trichrome (MT) and Picrosirius red (PSR) for circular polarized microscopy. Random quantitative analysis was performed on MT sections to classify the degree of IF, per field area, with and without circular polarization. Statistics correlations were performed by Spearman's (ρ; p < .05). There was a significant correlation of IF quantification with the area of interstitial collagen deposition by polarized PSR (PSRp) (r = .7939, p = .0098) and nonpolarized PSR (PSRn) (r = .7781, p = .0080). There was a positive correlation of serum creatinine (sCr) at different stages of CKD with PSRp (r = .7939, p = .0098), PSRn (r = .8667, p = .0027) and MT (r = .7818, p = .0117). Correlations between the percentage of quantified area was also positive from PSRp to PSRn (r = .9030, p = .0009) and PSRp to MT (r = .7939, p = .0098). The PSRN was also correlated with MT (r = .9273, p = .0001). The correlation with IF and sCr follows the disease evolution and the quantification of collagen by PSR is an excellent tool for analyzing the disease severity at different stages.

Different associations of cryoprotectants for testicular tissue of prepubertal cats submitted to vitrification.

The cryopreservation of testicular tissue is presented as the only alternative for the preservation of genetic material from prepubertal animals. However, this biotechnology is still being tested. The objective of this study was to evaluate the effect of different associations of cryoprotectants and the potential of cell proliferation after vitrification of testicular tissue of prepubertal cats. Five testicular pairs from five prepubertal cats were used, and each pair was divided into four fragments. Of these, one fragment composed of the control group (CG) and the rest were distributed in experimental groups according to the associations of cryoprotectants to be tested (dimethyl sulphoxide (DMSO)/glycerol (GLY); ethylene glycol (EG)/GLY) or DMSO/EG) in a final cryoprotectant concentration of 5.6 m. The fragments were submitted to vitrification, and after one week, fragments were heated and processed for histomorphological evaluation and quantification of nucleolar organizer regions (NORs). DMSO/GLY did not differ from CG and was superior to the other vitrified groups, as to cell separation and degree of shrinkage of the basal membrane. Concerning cell differentiation, visibility of the nucleus and nuclear condensation, all the vitrified groups were inferior to CG; however, DMSO/EG was inferior to DMSO/GLY and EG/GLY, which did not differ among themselves. CG was superior to all groups in quantification of NORs. DMSO/EG was inferior to all others, and there was no difference between DMSO/GLY and EG/GLY. The association DMSO/GLY presented the best preservation of tissue integrity and potential of cell proliferation after vitrification of the testicular tissue of prepubertal cats.

Two methods of vitrification followed by in vitro culture of the ovine ovary: evaluation of the follicular development and ovarian extracellular matrix.

The aim of this study was to evaluate the influence of two vitrification techniques on the extra cellular matrix (ECM) and ovarian follicular development. The ovarian cortex was fragmented (9 mm(3)) and divided into six groups, viz. fresh control, cultured control, vitrified by the Ovarian Tissue Cryosystem (OTC) method, conventional solid surface vitrification (SSV) method, OTC/cultured and SSV/cultured. Follicles from all the fragments were analysed for morphology, development and viability. The ECM was evaluated based on the condition of collagen and reticular fibres and the immunolocalization of type I collagen and fibronectin. After 7 days of culture, the tissue vitrified by OTC revealed a higher percentage (p < 0.05) of morphologically normal (30.66%) and viable (60.00%) follicles when compared with those vitrified using the SSV technique (21.33% and 23.00%). In all the fragments cultured, regardless of the vitrification method, a significantly higher percentage of developing follicles was observed when compared with the non-cultured tissue. Analysis of the type I collagen showed increased immunostaining after the in vitro culture in the vitrified fragments. In conclusion, the OTC is better for preserving the follicular viability and morphology and maintaining the integrity of the extracellular matrix components of the ovine ovary.

Screening of variables influencing the clavulanic acid production by Streptomyces DAUFPE 3060 strain.

Clavulanic acid (CA) is a beta-lactam antibiotic, which has a potent beta-lactamase inhibiting activity. The influence of five variables, namely pH (6.0, 6.4, and 6.8), temperature (28 degrees C, 30 degrees C, and 32 degrees C), agitation intensity (150, 200, and 250 rpm), glycerol concentration (5.0, 7.5, and 10 g/L) and soybean flour concentration (5.0, 12.5, and 20 g/L), on CA production by a new isolate of Streptomyces (DAUFPE 3060) was investigated in 250-mL Erlenmeyer flasks using a fractional factorial design. Temperature and soybean flour concentration were shown to be the two variables that exerted the most important effects on the production of CA at 95% confidence level. The highest CA concentration (494 mg/L) was obtained after 48 h at 150 rpm, 32 degrees C, pH 6.0, 5.0 g/L glycerol, and 20 g/L soybean flour concentrations. Under these conditions, the yields of biomass and product on consumed substrate were 0.26 g(X)/g(S) and 64.3 mg(P)/g(S), respectively. Fermentations performed in 3.0-L bench-scale fermenter allowed increasing the CA production by about 60%.

Intraflagellar transport complex in Leishmania spp. In silico genome-wide screening and annotation of gene function.

Flagella are constructed and maintained through the highly conserved process of intraflagellar transport (IFT), which is a rapid movement of particles along the axonemal microtubules of cilia/flagella. Particles that are transported by IFT are composed of several protein subunits comprising two complexes (A and B), which are conserved among green algae, nematodes, and vertebrates. To determine whether or not homologues to members of the IFT complex proteins are conserved in Leishmania spp, we scanned genomes, transcriptomes and proteomes of Leishmania species in a search for putative IFT factors, which were then identified in silico, compared, cataloged, and characterized. Since a large proportion of newly identified genes in L. major remain unclassified, with many of these being potentially Leishmania- (or kinetoplastid-) specific, there is a need for detailed analyses of homologs/orthologs that could help us understand the functional assignment of these gene products. We used a combination of integrated bioinformatics tools in a pathogenomics approach to contribute to the annotation of Leishmania genomes, particularly regarding flagellar genes and their roles in pathogenesis. This resulted in the formal in silico identification of eight of these homologs in Leishmania (IFT subunits, 20, 27, 46, 52, 57, 88, 140, and 172), along with others (IFTs 71, 74/72, and 81), as well as sequence comparisons and structural predictions. IFT, an important flagellar pathway in Leishmania, begins to be revealed through screening of trypanosomatid genomes; this information could also be used to better understand fundamental processes in Leishmania, such as motility and pathogenesis.

Intraflagellar transport complex in Leishmania spp. In silico genome-wide screening and annotation of gene function

Flagella are constructed and maintained through the highly conserved process of intraflagellar transport (IFT), which is a rapid movement of particles along the axonemal microtubules of cilia/flagella. Particles that are transported by IFT are composed of several protein subunits comprising two complexes (A and B), which are conserved among green algae, nematodes, and vertebrates. To determine whether or not homologues to members of the IFT complex proteins are conserved in Leishmania spp, we scanned genomes, transcriptomes and proteomes of Leishmania species in a search for putative IFT factors, which were then identified in silico, compared, cataloged, and characterized. Since a large proportion of newly identified genes in L. major remain unclassified, with many of these being potentially Leishmania- (or kinetoplastid-) specific, there is a need for detailed analyses of homologs/orthologs that could help us understand the functional assignment of these gene products. We used a combination of integrated bioinformatics tools in a pathogenomics approach to contribute to the annotation of Leishmania genomes, particularly regarding flagellar genes and their roles in pathogenesis. This resulted in the formal in silico identification of eight of these homologs in Leishmania (IFT subunits, 20, 27, 46, 52, 57, 88, 140, and 172), along with others (IFTs 71, 74/72, and 81), as well as sequence comparisons and structural predictions. IFT, an important flagellar pathway in Leishmania, begins to be revealed through screening of trypanosomatid genomes; this information could also be used to better understand fundamental processes in Leishmania, such as motility and pathogenesis.