Differential expression of inflammatory cytokines, prostaglandin synthases and secretory leukocyte protease inhibitor in the endometrium and circulation in different graded CEH-pyometra in bitch.

Cystic endometrial hyperplasia (CEH)-pyometra (CEH-P) is one of the most common reproductive disorders in bitches, posing a risk to both future fertility and life. The aims of the current study were to elucidate the differential expression patterns of inflammatory mediators at transcript and protein levels in the endometrium and to assess the concentrations of key inflammatory mediators in the peripheral circulation of bitches with different graded CEH-P. A total of 25 client-owned intact mixed breed bitches of 3-10 years presented to the outpatient department of RVP-TVCC of the institute were considered for the study. Of which, 22 cases suggestive of pyometra and 3 cases of CEH obtained during routine elective ovariohysterectomy were subjected to histopathological examination. Uteri were categorized into CEH (n = 3), moderate CEH-P (mCEH-P, n = 9), severe CEH-P (sCEH-P, n = 6) and atrophic pyometra (AT-P, n = 7). A group of age matched (n = 12) bitches without pyometra served as control. Endometrial transcripts such as IL6, IL8, PTGS2, PGFS, and SLPI were expressed differentially in the CEH and CEH-P bitch. In addition, a strong immunoreactivity (IR) of IL6, IL8, PTGS2, and mPGES1 was recorded in the sCEH-P uterus, while expression of IL10 was noticed in AT-P. In circulation, serum IL6 was the most relevant marker with high sensitivity of 96.2% and specificity of 84.6% at a cut off concentration 8.5 pg/mL followed by SLPI with 95.2% sensitivity, and 84.6% specificity at cut off concentration of 1.3 ng/mL. Serum IL10, PGFM and SLPI concentration in the peripheral circulation were 1.5-2.23 fold higher in mCEH-P, 0.87-2.5 fold higher in sCEH-P and 2.9-3.5 fold higher in AT-P than that of control. It is concluded that monitoring the serum concentration of IL6, IL10 and SLPI would be useful adjunct to the established hematobiochemical parameters in the management of pyometra in the bitch with critical illness.

Percutaneous transplantation of allogenic bone marrow-derived mesenchymal stem cells for the management of paraplegia secondary to Hansen type I intervertebral disc herniation in a Beagle dog.

BACKGROUND: Intervertebral disc herniation (IVDH) is one of the common causes of spinal cord injury (SCI) in dogs. It is commonly treated by performing surgical decompression that involves the removal of the extruded disc material. However, the recovery rates after surgical interventions are variable and many times unsatisfactory. This report aims to document a case of paraplegia associated with IVDH in a Beagle dog and its therapeutic management using allogenic bone marrow-derived mesenchymal stem cells (aBM-MSCs). CASE DESCRIPTION: The dog was presented with paraplegia that was initiated three weeks back. Based on the findings of computed tomography (CT), the condition was diagnosed as Hansen type I IVDH at T12-T13 intervertebral space. FINDINGS/TREATMENT AND OUTCOME: Neurological examination was performed to grade the neurological deficit. The isolation, culture, and characterization of aBM-MSCs were done as per the standard protocol. The prepared cell suspension of aBM-MSCs was percutaneously transplanted to the spinal cord parenchyma at the site of injury. A total of four doses of 1 × 106 cells were given at an interval of 15 days along with methylcobalamin and gabapentin orally. Improvement was evaluated based on the neurological examination and grading. Considerable improvement was noticed after the first dose of aBM-MSCs. The animal started complete weight bearing on its pelvic limbs after two doses. CONCLUSION: Percutaneous transplantation of aBM-MSCs might have played an important role in reversing the neurological deficits secondary to IVDH in this dog. Further studies are required preferably in a larger population to confirm the efficacy of aBM-MSCs therapy in ameliorating neural deficits associated with IVDH.

Diagnosis and surgical management of an intraocular foreign body secondary to ballistic wound in a Rhesus macaque (Macaca mulatta).

BACKGROUND: Intraocular foreign bodies (IOFBs) such as air gun pellet is a rare finding in wild animals like Rhesus macaque (Macaca mulatta). The purpose of the present scientific report is to describe the surgical retrieval of IOFB secondary to ballistic wound in a wild Rhesus macaque. CASE DESCRIPTION: A juvenile female wild Rhesus macaque was brought with the history of swollen and inflamed right eye for the past several days. FINDINGS/TREATMENT AND OUTCOME: Clinical examination revealed presence of partially healed wound over the dorsal eyelid. Radiographic examination revealed the presence of a metallic foreign body inside the right orbit. Inflamed and persistently closed eyelid prevented the further localization of the metallic foreign body. Lateral canthotomy was performed under general anesthesia. Following the failure to recover the metallic foreign body from the ocular adnexa, right eye vitrectomy was performed to retrieve the IOFB. The metallic foreign body was recovered from the posterior chamber of the right eye. Due to the poor prognosis of the already damaged eye, enucleation of the eye was performed which was followed by tarsorrhaphy. Further examination of the foreign body identified it as a 4.5 mm (.177 Calibre) air gun pellet. Post-operatively animal was treated with antibiotics and anti-inflammatory drugs. The animal recovered uneventfully. CONCLUSION: Intraocular foreign bodies secondary to gunshot wound should always be considered as a surgical emergency. Enucleation should be performed in cases having poor prognosis to avoid further complications especially in wild animals like Rhesus macaque.

In vitro proliferation and differentiation of canine bone marrow derived mesenchymal stem cells over hydroxyl functionalized CNT substrates.

Nanotopography of culture substrate acts as a positive cue in cell-biomaterial based tissue regeneration. Considering the potentiality of carbon nanotubes (CNTs) this study was designed to evaluate its two functionalized form by an in vitro culture condition using canine mesenchymal stem cells as cellular model. Cells were isolated and its behaviour, proliferation and differentiation processes were elucidated onto CNT substrates. Beside the variations in cellular behaviour it was remarkably noted that even though proliferation was reduced but osteogenic and chondrogenic differentiation was enhanced over multi-walled CNTs, whereas neuronal differentiation was better supported by single walled CNTs as evidenced by our cytochemical, immunocytochemical, gene expression and flow cytometry assays. The former one was noticed more cytocompatible by our different apoptosis studies. The outcome of these experiments collectively indicated that hydroxylated functionalized CNTs could be a potential scaffold constituent for future experimentations as well as for the application in regenerative medicine.

Preparation of NGF encapsulated chitosan nanoparticles and its evaluation on neuronal differentiation potentiality of canine mesenchymal stem cells.

Sustained and controlled release of neurotrophic factors in target tissue through nanomaterial based delivery system could be a better strategy for nerve tissue regeneration. The present study aims to prepare the nerve growth factor (NGF) encapsulated chitosan nanoparticles (NGF-CNPs) and its evaluation on neuronal differentiation potentiality of canine bone marrow derived mesenchymal stem cells (cBM-MSCs). The NGF-CNPs were prepared by ionotropic gelation method with tripolyphosphate (TPP) as an ionic cross-linking agent. Observations on physiochemical properties displayed the size of nanoparticles as 80-90 nm with positive zeta potential as well as an ionic interaction between NGF and nanoparticle. NGF loading efficiency was found to be 61% while its sustained release was observed by an in vitro release kinetics study. These nanoparticles were found to be cytocompatible to cBM-MSCs when supplemented at a concentration upto 4 mg/ml in culture media. The NGF-CNP supplemented culture media was able to transdifferentiate the preinduced cBM-MSCs into neurons in a better way than unbound NGF supplementation. Further, it was also noticed that NGF-CNPs were able to transdifferentiate cBM-MSCs without any chemical based preinduction. In conclusion, our findings propose that NGF-CNPs are capable of releasing bioactive NGF with the ability to transdifferentiate mesenchymal stem cells into neurons, suggesting its potential future application in nerve tissue regeneration.

Functionalized carbon nanotubes as suitable scaffold materials for proliferation and differentiation of canine mesenchymal stem cells.

In the field of regenerative medicine, numerous potential applications of mesenchymal stem cells (MSCs) can be envisaged, due to their ability to differentiate into a range of tissues on the basis of the substrate on which they grow. With the advances in nanotechnology, carbon nanotubes (CNTs) have been widely explored for use as cell culture substrate in tissue engineering applications. In this study, canine bone marrow-derived MSCs were considered as the cellular model for an in vitro study to elucidate the collective cellular processes, using three different varieties of thin films of functionalized carbon nanotubes (COOH-single-walled CNTs [SWCNTs], COOH-multiwalled CNTs [MWCNTs] and polyethylene glycol [PEG]-SWCNTs), which were spray dried onto preheated cover slips. Cells spread out better on the CNT films, resulting in higher cell surface area and occurrence of filopodia, with parallel orientation of stress fiber bundles. Canine MSCs proliferated at a slower rate on all types of CNT substrates compared to the control, but no decline in cell number was noticed during the study period. Expression of apoptosis-associated genes decreased on the CNT substrates as time progressed. On flow cytometry after AnnexinV-fluorescein isothiocyanate/propidium iodide (PI) staining, total number of apoptotic and necrotic cells remained lower in COOH-functionalized films compared to PEG-functionalized ones. Collectively, these results indicate that COOH-MWCNT substrate provided an environment of low cytotoxicity. Canine MSCs were further induced to differentiate along osteogenic, chondrogenic, and neuronal lineages by culturing under specific differentiation conditions. The cytochemical and immunocytochemical staining results, as well as the expression of the bone marker genes, led us to hypothesize that the COOH-MWCNT substrate acted as a better cue, accelerating the osteogenic differentiation process. However, while chondrogenesis was promoted by COOH-SWCNT, neuronal differentiation was promoted by both COOH-SWNCT and COOH-MWCNT. Taken together, these findings suggest that COOH-functionalized CNTs represent a promising scaffold component for future utilization in the selective differentiation of canine MSCs in regenerative medicine.

Caesarean in mare by Marcenac incision under local anaesthesia.

A nulliparous non-descript mare was presented with a complaint of dystocia. The mare was recumbent and physical examination revealed that the animal was in shock. There was no straining and foetal forelimbs were visible outside the vulva. The foetus was dead as there was no pedal reflex. Vaginal examination revealed anterior presentation with dorso-sacral position and rigid lateral head deviation. Pre-operatively, the mare was given 5 ml Tetanus toxoid and 3 g Ceftriaxone as intramuscular injection, and 5 ml Dexamethasone in 15 L of 5% Dextrose Normal Saline (DNS) as intravenous (i/v) infusion. As pelvic space was inadequate and the mal posture was not correctable, manual correction or foetotomy could not be attempted and therefore caesarean section was planned. Condition of the animal warranted the use of local anaesthetic infiltration instead of general anesthesia. Post-operative care included intravenous fluids, anti-inflammatory/analgesics and daily antiseptic dressing. The owner reported uneventful recovery. The authors would like to conclude the case as a rare emergency caesarean in equine where the surgery was done with animal in lateral recumbency employing a Marcenac incision under local anaesthesia.

Reference values of M-mode echocardiographic parameters and indices in conscious Labrador Retriever dogs.

Breed-wise standard echocardiographic values in dogs have been reported as there is variation in body and chest conformation which limits the application of data of one breed for other breed. Labrador Retrievers being originated from hunting dogs, might have different echocardiographic values from standard normal range of other dog breeds. So, the present study was aimed to determine the M-mode echocardiographic reference ranges in Labrador Retriever dogs and to evaluate the effect of body weight and gender on these parameters. The data obtained were also compared with that of the other dog breeds. Conscious clinically healthy Labrador Retriever dogs (n=24) of both sexes were made the subject of the study. All the measurements were made from a right parasternal long axis left ventricular outflow tract view and the parameters measured were: left ventricular dimensions, left ventricular function, left ventricular volumes, left atrial and aortic root diameter and mitral valve parameters. Data obtained were also compared with that available for other dog breeds. Significant correlation (P<0.05) with body weight was obtained for some of the left ventricular, atrial and mitral valve parameters, being strong for LAD, AOD, LVIDd, LVIDs, IVSd and IVSs (r>0.5); moderate for LVPWd, LVPWs, EPSS, EF Slope and SV (r=0.3 to 0.5); weak for EDV and ESV (r<0.3). Non-significant effect of gender was seen on all the echocardiographic parameters. However, some of the parameters had a significant breed effect. It is expected that the obtained data will be valuable for the progress of studies on small animal cardiology.

Using acellular aortic matrix to repair umbilical hernias of calves.

BACKGROUND: Umbilical hernias are a relatively common, possibly hereditary, condition in calves. Acellular aortic matrix (AAM) was evaluated for the repair of umbilical hernias in nine calves. METHODS: Fresh buffalo aorta to be used as a graft was decellularised using 1% sodium dodecyl sulfate and 0.25% trypsin. Under xylazine sedation and local analgesia, the hernial ring was exposed and repaired with the AAM graft using an inlay technique. RESULTS: All animals had an uneventful recovery without clinical signs of wound dehiscence, infection or recurrence of hernias during the 6-month follow-up period. CONCLUSION: AAM of buffalo origin has adequate strength to be used safely for the repair of umbilical hernias in calves.

Pancreatic beta-cell regeneration

Cephalendra indica (41//v/v alcoholic extract of the wild variety of Cephalendra indica Naud.), on regular administration in doses ranging from 25uml to 75uml/100g of body weight (gbw) by the oral or intraperitoneal (ip) route produced a significant fall in blood sugar level in alloxan-induced diabetic rats. Biochemical studies showed stabilization of blood sugar level in 70//of cases at fourteen to twenty days after withdrawal of the drug. Histopathological studies revealed regeneration of pancreatic B cells. The hypothesis is that the drug acts through the hypothalamo-hypophysial-pancreatic axis, producing selective regeneration of B cells. The drug may indirectly release inhibitory factors from hypothalamic neurons, inhibiting the secretion of growth hormone and triggering insulin secretions from B cells. The therapeutic action of the drug on pancreatic B cells and lack of acute and subacute toxicity may open up new prospects in the treatment of diabetes mellitus