Aberrant expression of microRNAs and the miR-1/MET pathway in canine hepatocellular carcinoma.

Canine hepatocellular carcinoma (HCC) is the most common primary hepatic tumour in dogs. MicroRNA (miRNA) dysregulation has been reported in human HCC and shown to have diagnostic and prognostic value; however, there are no data on miRNA expression in canine HCC. The aim of the present study was to investigate differentially expressed miRNAs in canine HCC. Analysis of miRNA expression in canine HCC tissues and cell lines by quantitative reverse transcription PCR showed that miR-1, miR-122, let-7a, and let-7g were downregulated, whereas miR-10b and miR-21 were upregulated in canine HCC. MET is one of the target genes of miR-1. MET was upregulated in canine HCC at the gene and protein levels, and a significant correlation between the concomitant downregulation of miR-1 and upregulation of MET was observed. Fast/intermediate-proliferating canine HCC cell lines had higher MET gene and protein expression levels than the slow-proliferating cell line. These findings suggest that miRNAs are differentially expressed in canine HCC, and that the miR-1/MET pathway may be associated with canine HCC cell proliferation.

Longitudinal and long-term effects of radiotherapy on swallowing function after tongue reconstruction.

OBJECTIVE: This study evaluated the longitudinal and long-term effects of radiotherapy on swallowing function after tongue reconstruction. METHODS: The study comprised 16 patients who had: undergone glossectomy and tongue reconstruction with free flap transfer, received adjuvant radiotherapy, and survived without recurrence for at least 1 year. Swallowing function, as indicated by tolerance of oral intake, was evaluated before radiotherapy, at radiotherapy completion, and at 6 and 12 months after radiotherapy completion. RESULTS: Before radiotherapy, all patients could tolerate oral intake. At radiotherapy completion, only three patients could consume all nutrition orally. However, swallowing function improved over time, and by 12 months after radiotherapy completion it had returned nearly to that before radiotherapy. CONCLUSION: Acute dysphagia due to radiotherapy after tongue reconstruction is severe, but can improve gradually. Multidisciplinary support of patients during percutaneous endoscopic gastrostomy dependence is important to improve long-term functional outcomes.

En Bloc Multivisceral and Kidney Transplantation in an HIV Patient: First Case Report.

The continual improvement in outcome with highly active antiretroviral therapy (HAART) for human immunodeficiency virus (HIV) infection and visceral transplantation for gut failure stimulated our interest in lifting HIV infection as a contraindication for intestinal and multivisceral transplantation. This report is the first to describe visceral transplantation in a patient with HIV infection. A HAART regimen was introduced in the setting of short-gut syndrome with successful suppression of HIV viral load. The indication for en bloc multivisceral and kidney transplantation was end-stage liver failure with portomesenteric venous thrombosis and chronic renal insufficiency. The underlying hepatic pathology was alcoholic and home parenteral nutrition-associated cirrhosis. Surgery was complicated due to technical difficulties with excessive blood loss and long operative time. The complex posttransplant course included multiple exploratory laparotomies due to serious intra-abdominal and systemic infections. Heavy immunosuppression was required to treat recurrent episodes of severe allograft rejection. Posttransplant oral HAART successfully sustained undetectable viral load. Unfortunately, the patient succumbed to sepsis 3 months posttransplant. With new insights into the biology of gut immunity, mechanisms of allograft tolerance, and HIV-associated immune dysregulation, successful outcome is anticipated, particularly in patients who are in need of isolated intestinal and less-organ-contained visceral allografts.

Multipotency of equine mesenchymal stem cells derived from synovial fluid.

Cartilage regeneration with cell therapy following arthroscopic surgery could be used in racehorses with intra-articular fractures (IAF) and osteochondritis dissecans (OCD). The aims of this study were to investigate the origin and multipotency of stromal cells in the synovial fluid (SF) of horses with intra-articular injury and synovitis, and to provide a new strategy for regeneration of lost articular cartilage. Mesenchymal stromal cells were isolated from SF of horses with IAF and OCD. Multipotency was analysed by RT-PCR for specific mRNAs and staining for production of specific extracellular matrices after induction of differentiation. The total number of SF-derived mesenchymal stromal cells reached >1 × 10(7) by the fourth passage. SF-derived cells were strongly positive (>90% cells positive) for CD44, CD90 and major histocompatibility complex (MHC) class I, and moderately positive (60-80% cells positive) for CD11a/CD18, CD105 and MHC class II by flow cytometry. SF-derived cells were negative for CD34 and CD45. Under specific nutrient conditions, SF-derived cells differentiated into osteogenic, chondrogenic, adipogenic and tenogenic lineages, as indicated by the expression of specific marker genes and by the production of specific extracellular matrices. Chondrogenic induction in culture resulted in a change in cell shape to a 'stone-wall' appearance and formation of a gelatinous sheet that was intensely stained with Alcian blue. SF may be a novel source of multipotent mesenchymal stem cells with the ability to regenerate chondrocytes.

Split liver transplantation using Hemiliver graft in the MELD era: a single center experience in the United States.

Under the "sickest first" Model for End-Stage Liver Disease (MELD) allocation, livers amenable to splitting are most often allocated to patients unsuitable for split liver transplantation (SLT). Our experience with SLT using hemilivers was reviewed. From April 2004 to June 2012, we used 25 lobar grafts (10 left lobes and 15 right lobes) for adult-sized recipients. Twelve recipients were transplanted with primary offers, and 13 were transplanted with leftover grafts. Six grafts were shared with other centers. The data were compared with matched whole liver grafts (n = 121). In 92% of donors, the livers were split in situ. Hemiliver recipients with severe portal hypertension had a greater graft-to-recipient weight ratio than those without severe portal hypertension (1.96% vs. 1.40%, p < 0.05). Hemiliver recipients experienced biliary complications more frequently (32.0% vs. 10.7%, p = 0.01); however, the 5-year graft survival for hemilivers was comparable to whole livers (80.0% vs. 81.5%, p = 0.43). The secondary recipients with leftover grafts did not have increased incidences of graft failure (p = 0.99) or surgical complications (p = 0.43) compared to the primary recipients. In conclusion, while routine application is still controversial due to various challenges, hemiliver SLT can achieve excellent outcomes under the MELD allocation.

Comparison of multiple parameters obtained on 3T pulsed arterial spin-labeling, diffusion tensor imaging, and MRS and the Ki-67 labeling index in evaluating glioma grading.

BACKGROUND AND PURPOSE: Pulsed arterial spin-labeling, DTI, and MR spectroscopy provide useful data for tumor evaluation. We evaluated multiple parameters by using these pulse sequences and the Ki-67 labeling index in newly diagnosed supratentorial gliomas. MATERIALS AND METHODS: All 32 patients, with grade II (3 each of diffuse astrocytoma, oligodendroglioma, and oligoastrocytoma), grade III (3 anaplastic astrocytomas, 4 anaplastic oligodendrogliomas, and 1 anaplastic oligoastrocytoma), and grade IV (14 glioblastomas and 1 glioblastoma with an oligodendroglioma component) cases underwent pulsed arterial spin-labeling, DTI, and MR spectroscopy studies by using 3T MR imaging. The following variables were used to compare the tumors: relative cerebral blood flow, fractional anisotropy; ADC tumor/normal ratios; and the Cho/Cr, NAA/Cho, NAA/Cr, and lactate/Cr ratios. A logistic regression and receiver operating characteristic analysis were used to assess parameters with a high sensitivity and specificity to identify the threshold values for separate grading. We compared the Ki-67 index with various MR imaging parameters in tumor specimens. RESULTS: Significant correlations were observed between the Ki-67 index and the mean, maximum, and minimum ADC, Cho/Cr, and lactate/Cr ratios. The receiver operating characteristic analysis showed that the combination of the minimum ADC and Cho/Cr ratios could differentiate low-grade and high-grade gliomas, with a sensitivity and specificity of 87.0% and 88.9%, respectively. The mean and maximum relative cerebral blood flow ratios were used to classify glioblastomas from other-grade astrocytomas, with a sensitivity and specificity of 92.9% and 83.3%, respectively. CONCLUSIONS: Our findings indicate that pulsed arterial spin-labeling, DTI, and MR spectroscopy are useful for predicting glioma grade. Additionally, the parameters obtained on DTI and MR spectroscopy closely correlated with the proliferative potential of gliomas.

The clinical and pathological significance of nitric oxide synthase in human pituitary adenomas: a comparison with MIB-1.

The purpose of the present study was to define the clinical and pathological significance of nitric oxide synthase (NOS) in human pituitary adenomas, and to compare these values with those of the MIB-1 labeling index (LI) using an immunohistochemical method. Tissue specimens from 82 cases of surgically-treated pituitary adenomas were immunostained for hormone production for the MIB-1 LI and for the three NOS isoenzymes and five normal pituitary glands were immunostained for the three NOS isoenzymes as a control. The correlation between the clinical variables (age, functional status, tumor size, Hardy's grading, cavernous and/or sphenoid invasiveness, and progression) and mean MIB-1 LI, or between the same clinical variables and NOS immunoreactivity (IR) were analyzed. There was a statistically significant difference in the MIB-1 LI between macroadenomas and microadenomas, and between invasive adenomas and noninvasive adenomas. On the other hand, there was a statistically significant difference in the inducible NOS (iNOS) IR between invasive adenomas and noninvasive adenomas. Furthermore, the iNOS IR had a significant correlation with the MIB-1 LI. Invasive adenomas have a higher iNOS IR, and this correlated with the MIB-1 LI. These findings may be due to the function of iNOS, which plays an important role in tissue injury and repair.

Adjuvant treatment of hepatocellular carcinoma after orthotopic liver transplantation: do we really need this?

Hepatocellular carcinoma (HCC) continues to rise and is still a major cause of mortality. Orthotopic liver transplantation (OLT) continues to give patients the best chance for cure, but recurrence of the disease remains a problem. Even with the implementation of the Milan criteria, recurrence rates have been shown to be 8-15% in most studies and even higher in patients who are beyond the Milan criteria. Therefore, several investigators have looked into the value of adjuvant therapy using systemic cytotoxic chemotherapy in HCC after OLT. Unfortunately, most of the trials are very small, and the results have been disappointing. But trials using Licartin seem to be promising, and other drugs such as FOLFOX and sorafenib warrant further investigation based on their efficacy in the advanced disease. In this review, we will review the current data on efficacy and rationale of adjuvant treatment for HCC after OLT including novel biomarkers.

Matrix metalloprotease-9 activity in the cerebrospinal fluid and spinal injury severity in dogs with intervertebral disc herniation.

We investigated whether matrix metalloproteinase (MMP)-9 expression in the cerebrospinal fluid (CSF) of dogs with intervertebral disc herniation (IVDH) is associated with the severity of neurological signs and prognosis. CSF from the cisterna magna (C-CSF) and the lumbar spine (L-CSF) of 34 dogs with IVDH was analyzed using zymography. Activity of MMP-9 in L-CSF was detected in 6 of 34 dogs with IVDH, often for more than 7 days after injury. MMP-9 activity was not detected from any of the C-CSF samples. Of the six cases that were MMP-9 positive, all four cases with grade V that had loss of deep pain were non-ambulatory 6 months after treatment. The remaining two cases with grade III and IV could recover mobility. In dogs with grade V thoracolumbar IVDH, MMP-9 expression in the CSF may indicate severe spinal cord injury with poor prognosis.

Use of tissue plasminogen activator in liver transplantation from donation after cardiac death donors.

Ischemic-type biliary stricture (ITBS) occurs in up to 50% after liver transplantation (LT) from donation after cardiac death (DCD) donors. Thrombus formation in the peribiliary microcirculation is a postulated mechanism. The aim was to describe our experience of tissue plasminogen activator (TPA) administration in DCD-LT. TPA was injected into the donor hepatic artery on the backtable (n = 22). Two recipients developed ITBS including one graft failure. Although excessive postreperfusion bleeding was seen in 14 recipients, the amount of TPA was comparable between those with and without excessive bleeding (6.4 ± 2.8 vs. 6.6 ± 2.8 mg, p = 0.78). However, donor age (41 ± 12 vs. 29 ± 9 years, p = 0.02), donor BMI (26.3 ± 5.5 vs. 21.7 ± 3.6 kg/m(2) , p = 0.03), previous laparotomy (50% vs. 0%, p = 0.02) and lactate after portal reperfusion (6.3 ± 4.6 vs. 2.8 ± 0.9 mmol/L, p = 0.005) were significantly greater in recipients with excessive bleeding. In conclusion, the use of TPA may lower the risk of ITBS-related graft failure in DCD-LT. Excessive bleeding may be related to poor graft quality and previous laparotomy rather than the amount of TPA. Further studies are needed in larger population.

Comparative study of chronic kidney disease in dogs and cats: induction of myofibroblasts.

We investigated the kidneys of dogs and cats to clarify whether renal myofibroblasts induction is associated with the severity of chronic kidney disease (CKD). Immunohistochemical expression of myofibroblast markers, alpha-smooth muscle actin (SMA) and vimentin, were evaluated quantitatively. The degrees of glomerulosclerosis, glomerular hypertrophy, interstitial cell infiltration, and interstitial fibrosis were also evaluated quantitatively. The plasma creatinine (pCre) concentrations correlated with glomerulosclerosis, cell infiltration, and fibrosis in dogs, and only with fibrosis in cats. The alpha-SMA expression correlated with pCre, glomerulosclerosis, cell infiltration, and fibrosis in dogs, and with pCre and fibrosis in cats. Tubular vimentin expression correlated with fibrosis in cats, but not in dogs. Interstitial vimentin expression correlated with pCre, glomerulosclerosis, cell infiltration, and fibrosis in dogs, but only with pCre in cats. In conclusion, it was suggested that the severity of CKD in dogs and cats was mediated by different pathways associated with myofibroblasts expression.

Significance of des-gamma-carboxy prothrombin in selection criteria for living donor liver transplantation for hepatocellular carcinoma.

Des-gamma-carboxy prothrombin (DCP) levels reportedly correlate with histological features of hepatocellular carcinoma (HCC). We examined serum DCP as a predictor of HCC recurrence in 144 patients who underwent living donor liver transplantation. Receiver operating characteristics (ROC) analysis revealed superiority of DCP and AFP over preoperative tumor size or number for predicting recurrence. Multivariate analysis revealed tumor size >5 cm, > or =11 nodules, and DCP >400 mAU/mL as significant independent risk factors for recurrence. Incidence of microvascular invasion (62% vs. 27%, p = 0.0003) and poor differentiation (38% vs. 16%, p = 0.0087) were significantly higher for patients with DCP >400 mAU/mL than for patients with DCP < or =400 mAU/mL. In ROC analysis for patients with < or =10 nodules all < or =5 cm to predict recurrence, area under the curve was much higher for DCP than for AFP (0.84 vs. 0.69). Kyoto criteria were thus defined as < or =10 nodules all < or =5 cm, and DCP < or =400 mAU/mL. The 5-year recurrence rate for 28 patients beyond-Milan but within-Kyoto criteria was as excellent as that for 78 patients within-Milan criteria (3% vs. 7%). The preoperative DCP level offers additional information regarding histological features, and thus can greatly improve patient selection criteria when used with tumor bulk information.

Immunohistochemical analysis of cyclooxygenase-2 induction during wound healing in dog skin.

Cyclooxygenase (COX)-2 is an inducible isoform of COX and is expressed under abnormal health conditions. This study elucidated the cutaneous induction of COX-2 during the wound healing processes in dog skin. Dog skin was sutured after punch biopsy and investigated histologically and immunohistochemically on days 0 (normal), 1, 3, 5, 7, 10, and 14 after injury. Histological changes, including infiltration of inflammatory cells and proliferation of fibroblast-like cells, were observed as predicted, and there was a close and significant correlation between these 2 events. COX-2-positive cells were detected in the epidermis between days 1 and 7, and bimodal peaks were observed in the case of the percentage of COX-2-positive cells. In inflammatory cells, COX-positive signals were detected on day 3 only. Here, we clarified the localization and pattern of the induced COX-2 expression during wound healing in dog skin.

The association between high VEGF levels and multiple probable punctuate cavernous malformations.

Cerebral cavernous malformations (CCMs) are congenital abnormalities of the cerebral vessels. The de novo development of new lesions in this disease has been reported. However, the underlying mechanism of progressive CCMs in such patients remains unclear. This report documents two cases of multiple probable CCMs that showed a progressive behaviour. The plasma levels of vascular endothelial growth factor (VEGF), and transforming growth factor-beta1 (TGF-beta1) were measured using an enzyme-linked immunosorbent assay (ELISA). The concentration of both VEGF and TGF-beta1 in plasma was increased in these patients. A relationship was observed between high concentrations of growth factors and progressive CCMs. Even though a causal linkage between these conditions cannot be confirmed, a continuous high VEGF level in plasma could be a possible clinical indicator for subsequent intracerebral haemorrhages in the CCM patients.

Neuroprotective effects of edaravone, a free radical scavenger, on the rat hippocampus after pilocarpine-induced status epilepticus.

PURPOSE: Edaravone (MCI-186) is a newly developed antioxidative radical scavenger for the treatment of acute cerebral infarction, exerting neuroprotective effects against ischemic insult. The neuroprotective effects of edaravone on pilocarpine-induced seizures in rats were investigated. METHODS: Rats were treated intraperitoneally with saline or edaravone (1-30 mg/kg), applied 30 min before pilocarpine hydrochloride (330 mg/kg). The onset of status epilepticus (SE) and mortality were recorded for a period of at least 3 days. The cell loss and immunoreactivities of nitric oxide synthase (NOS) in the hippocampus from control and the day 3 rats after SE, treated with saline or edaravone, were evaluated. RESULTS: Edaravone (1mg/kg) significantly prevented cell loss in the hippocampus after SE while easier inducing SE. The higher dose of drug could not induce SE significantly but tended to increase the rate of mortality. Inducible NOS (iNOS) expression was significantly decreased in the hippocampus from day 3 rats treated with 1mg/kg edaravone, compared with saline group, while neuronal NOS (nNOS) and iNOS significantly increased in the hippocampus treated with saline, compared with control group. Significant alteration of endothelial NOS (eNOS) expression in the hippocampus among control group, saline group, and edaravone group was not shown. CONCLUSIONS: Edaravone may act as a neuroprotector for the hippocampus after SE by reducing at least iNOS although the low dose of drug easier induces SE because of preventing an endogenous antiepileptic effect of NO.

Hyperhomocysteinaemia is a significant risk factor for white matter lesions in Japanese type 2 diabetic patients.

The presence of white matter lesions (WML) is an important prognostic factor for the development of stroke. Plasma total homocysteine (tHcy), which increases with diabetes, has been flagged as a novel predictor for cerebrovascular events. We tested the hypothesis that the presence of WML correlates with tHcy and insulin resistance in type 2 diabetic patients not receiving insulin treatment. Based on brain magnetic resonance imaging findings, 81 type 2 diabetic patients were divided into two groups, with-WML group (57 +/- 8 years, mean +/- standard deviation, n = 31) and without-WML group (57 +/- 6 years, n = 50). The blood glucose level was assessed by fasting plasma glucose (FPG), fasting immunoreactive insulin, Homeostasis Model Assessment (HOMA) Index and hemoglobin A1c. The body mass index was higher in the with-WML group than in the without-WML group (P < 0.05). Plasma levels of triglyceride were higher whilst high-density lipoprotein cholesterol was lower in the with-WML group than in the without-WML group (P < 0.05 and P < 0.0001 respectively). FPG (P < 0.005), insulin concentrations (P < 0.0001), HOMA Index (P < 0.0001) and tHcy (<0.0001) levels were higher in the with-WML group than in the without-WML group. Multivariate logistic analysis revealed that WML was independently predicted by the high tHcy and insulin resistance. Our findings indicate that the presence of WML was associated with the high tHcy and insulin resistance in these Japanese patients with type 2 diabetes mellitus.

Nitric oxide synthase and NMDA receptor expressions in cavernoma tissues with epileptogenesis.

OBJECTIVES: To investigate the contribution of nitric oxide (NO) and the glutamate systems to epileptogenicity of cavernoma (CA). METHODS: Using immunohistochemistry we examined NO synthases (NOS; neuronal, inducible and endothelial) and N-methyl-D-aspartate (NMDA) receptor subunits 1(NR1) and 2A/B (NR2A/B) in tissues, with and without hemosiderin deposits, adjacent to CA resected from temporal (seven patients) and frontal (one patient) lobes. RESULTS: All isoforms of NOS, especially iNOS expression, was significantly upregulated in company with NR2A/B expression, not only in declining neuronal cells but also in reactive astrocytes in the tissue, with hemosiderin deposits, adjacent to CA and moreover the degree of iNOS expression was significantly correlated with seizure frequency. CONCLUSIONS: These preliminary results sustain a speculation that excessive NO may generate in the tissue surrounding CA with repeated microhaemorrhaging and seizures. The neuronal loss and reactive glial proliferation induced by iron or NO may play a role in epileptogenesis.

Spontaneous chronic head and spinal subdural haematoma.

Spinal chronic subdural haematomas (SSDH) are extremely rare. We report a case of a SSDH combined with intracranial SDH. After tapping the SSDH in addition to the evacuation of SDH, the clinical symptoms dramatically improved. SSDH are considered to have an uncertain prognosis; however, good results can be obtained with an early diagnosis and prompt treatment.

Overlapping free bone graft with galea-pericranium in reconstruction of the anterior skull base to prevent CSF leak and sequestrum formation.

Background. Reconstruction of the skull base after resection of a tumour is important to prevent postoperative complications such as infectionsand cerebrospinal fluid (CSF) leakage. Several reconstructive methods of the anterior skull base have been reported but, their long-term results are not clear. Methods. We describe a technique used after removal of an olfactory neuroblastoma with infiltration of the skull base. The reconstructed dura was covered with a galeal patch, a replicated galeal-pericranial flap, a graft from the inner table of skull, and a vascularised galeal-pericranial flap placed on the skull base defect. All layers were fixed with fibrin glue. Conclusion. Three dimensional computed tomography (3D-CT) at bone window settings demonstrated the bone graft covered the bone defect and was not absorbed and after 11 years there have been no signs of tumour regrowth or complications.

The relationship between paired pulse magnetic MEP and surgical prognosis in patients with intractable epilepsy.

PURPOSE: To assess whether paired pulse magnetic motor evoked potential (MEP) can predict surgical prognosis in patients with intractable epilepsy. METHODS: MEP of the unilateral hand muscles were recorded following paired pulse transcranial magnetic stimulation (TMS) of the motor cortex. The interstimulus intervals of paired stimulation were 1-16 ms with a conditioning stimulus that was 90% active motor threshold. Subjects were six patients with temporal lobe epilepsy (TLE) scheduled for anterior temporal lobectomy and three patients with myoclonic or head-drop seizures scheduled for anterior corpus callosotomy, resulting in the unilateralization of epileptic discharges. The hemisphere showing unilateral discharges was defined as the affected hemisphere. The intracortical inhibition and facilitation curve was drawn based on MEP before and after surgery and the relationship between MEP and surgical prognosis was investigated. RESULTS: In five patients with TLE showing class I surgical results (Engel's classification), the affected hemisphere showing cortical hyperexcitability preoperatively was almost normalized after surgery. However, in a patient with class III, the unaffected hemisphere showed cortical hyperexcitability before and after surgery. In the callosotomy group, two patients with excellent outcomes showed the same results as TLE group with class I. CONCLUSIONS: Paired pulse magnetic MEP may provide predictive value in terms of surgical outcome in those patients with intractable epilepsy.