Preventive and Therapeutic Effects of Krill Oil on Obesity and Obesity-Induced Metabolic Syndromes in High-Fat Diet-Fed Mice.

Obesity increases the risks of metabolic syndromes including nonalcoholic fatty liver disease (NAFLD), diabetic dyslipidemia, and chronic kidney disease. Dietary krill oil (KO) has shown antioxidant and anti-inflammatory properties, thereby being a therapeutic potential for obesity-induced metabolic syndromes. Thus, the effects of KO on lipid metabolic alteration were examined in a high-fat diet (HFD)-fed mice model. The HFD model (n = 10 per group) received an oral gavage with distilled water as a control, metformin at 250 mg/kg, and KO at 400, 200, and 100 mg/kg for 12 weeks. The HFD-induced weight gain and fat deposition were significantly reduced in the KO treatments compared with the control. Blood levels were lower in parameters for NAFLD (e.g., alanine aminotransferase, and triglyceride), type 2 diabetes (e.g., glucose and insulin), and renal dysfunction (e.g., blood urea nitrogen and creatinine) by the KO treatments. The KO inhibited lipid synthesis through the modification of gene expressions in the liver and adipose tissues and adipokine-mediated pathways. Furthermore, KO showed hepatic antioxidant activities and glucose lowering effects. Histopathological analyses revealed that the KO ameliorated the hepatic steatosis, pancreatic endocrine/exocrine alteration, adipose tissue hypertrophy, and renal steatosis. These analyses suggest that KO may be promising for inhibiting obesity and metabolic syndromes.

Molecular Detection and Genetic Diversity of Blastocystis in Korean Dogs.

Blastocystis is a genus of unicellular heterokont parasites belonging to a group of organisms known as Stramenopiles, which includes algae, diatoms, and water molds. Blastocystis includes several species that habitat in the gastrointestinal tracts of organisms as diverse as humans, farm animals, birds, rodents, reptiles, amphibians, fish, and cockroaches. It is important to public health and distributed globally, but its prevalence in dogs in Korea has not been reported to date. Here, we collected 787 canine fecal samples and assessed Blastocystis infection by age, sex, region, season, and diarrhea symptoms. We determined Blastocystis subtypes using phylogenetic analyses based on 18S rRNA gene sequences. We identified, 10 Blastocystis positive samples (1.3%). A higher proportion of infected dogs was asymptomatic; however, infection rates did not significantly differ according to region, age, sex, and season. Phylogenetic analysis showed that the Blastocystis sp. identified belonged to 4 subtypes (STs), ST1, ST5, ST10, and ST14, thus revealed the genetic diversity of Blastocystis sp. in dogs Korean. This is first report on the presence of Blastocystis sp. in dogs Korean. This study revealed a lower infection rate than expected and differed from previous studies in STs. Further studies are warranted to observe the national infection status of Blastocystis in dogs and the genetic characteristics of this genus.

Cytoglobin deficiency potentiates Crb1-mediated retinal degeneration in rd8 mice.

PURPOSE: The purpose of this study is to determine the effect of Cytoglobin (Cygb) deficiency on Crb1-related retinopathy. The Crb1 cell polarity complex is required for photoreceptor function and survival. Crb1-related retinopathies encompass a broad range of phenotypes which are not completely explained by the variability of Crb1 mutations. Genes thought to modify Crb1 function are therefore important targets of research. The biological function of Cygb involves oxygen delivery, scavenging of reactive oxygen species, and nitric oxide metabolism. However, the relationship of Cygb to diseases involving the Crb1 cell polarity complex is unknown. METHODS: Cygb knockout mice homozygous for the rd8 mutation (Cygb-/-rd8/rd8) were screened for ocular abnormalities and imaged using optical coherence tomography and fundus photography. Electroretinography was performed, as was histology and immunohistochemistry. Quantitative PCR was used to determine the effect of Cygb deficiency on transcription of Crb1 related cell polarity genes. RESULTS: Cygb-/-rd8/rd8 mice develop an abnormal retina with severe lamination abnormalities. The retina undergoes progressive degeneration with the ventral retina more severely affected than the dorsal retina. Cygb expression is in neurons of the retinal ganglion cell layer and inner nuclear layer. Immunohistochemical studies suggest that cell death predominates in the photoreceptors. Electroretinography amplitudes show reduced a- and b-waves, consistent with photoreceptor disease. Cygb deficient retinas had only modest transcriptional perturbations of Crb1-related cell polarity genes. Cygb-/- mice without the rd8 mutation did not exhibit obvious retinal abnormalities. CONCLUSIONS: Cygb is necessary for retinal lamination, maintenance of cell polarity, and photoreceptor survival in rd8 mice. These results are consistent with Cygb as a disease modifying gene in Crb1-related retinopathy. Further studies are necessary to investigate the role of Cygb in the human retina.

Anti-Photoaging Effects of Low Molecular-Weight Fucoidan on Ultraviolet B-Irradiated Mice.

Ultraviolet (UV) B exposure induces DNA damage and production of reactive oxygen species (ROS), which causes skin photoaging through signaling pathways of inflammation and modulation of extracellular matrix remodeling proteins, collagens, and matrix metalloproteinase (MMP). As low molecular-weight fucoidan (LMF) has potential antioxidant and anti-inflammatory properties, we examined the protective effects of LMF against UVB-induced photoaging. A UVB-irradiated mouse model was topically treated with myricetin or LMF at 2.0, 1.0 and 0.2 mg/cm² (LMF2.0, LMF1.0 and LMF0.2, respectively) once a day for 15 weeks. Wrinkle formation, inflammation, oxidative stress, MMP expression, and apoptosis in the treated regions were compared with those in a distilled water-treated photoaging model (UVB control). LMF treatments, particularly LMF2.0 and LMF1.0, significantly inhibited the wrinkle formation, skin edema, and neutrophil recruitment into the photo-damaged lesions, compared with those in the UVB control. While LMF decreased interleukin (IL)-1ß release, it increased IL-10. The LMF treatment inhibited the oxidative stresses (malondialdehyde and superoxide anion) and enhanced endogenous antioxidants (glutathione). Additionally, LMF reduced the mRNA expression of MMP-1, 9, and 13. The histopathological analyses revealed the anti-photoaging effects of LMF exerted via its antioxidant, anti-apoptotic, and MMP-9-inhibiting effects. These suggest that LMF can be used as a skin-protective remedy for photoaging.

Effect of platelet-rich plasma in Achilles tendon allograft in rabbits.

BACKGROUND: Achilles tendon is composed of dense connective tissue and is one of the largest tendons in the body. In veterinary medicine, acute ruptures are associated with impact injury or sharp trauma. Healing of the ruptured tendon is challenging because of poor blood and nerve supply as well as the residual cell population. Platelet-rich plasma (PRP) contains numerous bioactive agents and growth factors and has been utilized to promote healing in bone, soft tissue, and tendons. OBJECTIVE: The purpose of this study was to evaluate the healing effect of PRP injected into the surrounding fascia of the Achilles tendon after allograft in rabbits. METHODS: Donor rabbits (n = 8) were anesthetized and 16 lateral gastrocnemius tendons were fully transected bilaterally. Transected tendons were decellularized and stored at -80°C prior to allograft. The allograft was placed on the partially transected medial gastrocnemius tendon in the left hindlimb of 16 rabbits. The allograft PRP group (n = 8) had 0.3 mL of PRP administered in the tendon and the allograft control group (n = 8) did not receive any treatment. After 8 weeks, rabbits were euthanatized and allograft tendons were transected for macroscopic, biomechanical, and histological assessment. RESULTS: The allograft PRP group exhibited superior macroscopic assessment scores, greater tensile strength, and a histologically enhanced healing process compared to those in the allograft control group. CONCLUSIONS: Our results suggest administration of PRP on an allograft tendon has a positive effect on the healing process in a ruptured Achilles tendon.

Decreased SMP30 Expression Is Related With EMT in the Kidneys of Two Siberian Tigers With CKD.

BACKGROUND/AIM: Chronic kidney disease (CKD) is one of the most common causes of mortality in wild non-domestic felidae. The molecular mechanism regulating renal fibrosis in nephropathy is not fully understood especially in the felidae. This study aimed to elucidate senescence marker protein 30 (SMP30) expression patterns and its relationship with epithelial-mesenchymal transition (EMT) by immunostaining in two necropsied Siberian tigers (Panthera tigris altaica) with CKD. MATERIALS AND METHODS: Two kidney samples from male Siberian tigers were fixed and tissue sections were stained for histopathological assay. RESULTS: In CKD, renal tubular epithelial cells lost their tubular structures surrounded by severe interstitial fibrosis and were detached from the basement membrane. These damaged cells resembled the morphology of mesenchymal cells and showed much lower SMP30 expression compared with intact tubular epithelial cells. These cells also expressed vimentin, which is specifically expressed by mesenchymal cells, and through double staining, it was observed that vimentin was expressed in the tubular epithelial cells where SMP30 was not expressed. In addition, double-positive expression of pan-cytokeratin (pan-CK) and vimentin was found in damaged epithelial cells with mesenchymal features. CONCLUSION: We demonstrated possible evidence to understand the role of SMP30 as a new pivotal factor and the possibility of decreased SMP30 as a potential indicator of EMT at the end stage of CKD.

Cardiovascular and Respiratory Effects of Increased Intra-Abdominal Pressure with and without Dexmedetomidine in Anesthetized Dogs.

Intra-abdominal pressure (IAP) elevation during capnoperitoneum can cause adverse cardiovascular and respiratory effects. This study aimed to determine if a sequentially increased IAP affects cardiovascular and respiratory variables in anesthetized dogs and evaluate the effects of the constant-rate infusion of dexmedetomidine (Dex) on cardiovascular and respiratory variables with increased IAP. Five dogs were anesthetized and instrumented, and a Veress needle was equipped to adjust the IAP using a carbon dioxide insufflator. Stabilization was conducted for 1 h, and physiological variables were measured at IAPs of 0, 5, 10, 15, and 20 mmHg and after desufflation. After the washout period, the dogs underwent similar procedures along with a constant-rate infusion of dexmedetomidine. The cardiovascular effects of increased IAP up to 20 mmHg were not significant in healthy beagle dogs and those administered with dexmedetomidine. When comparing the control and dexmedetomidine groups, the overall significant effects of dexmedetomidine were noted on heart rate, cardiac output, and systemic vascular resistance during the experiment. Respiratory effects were not observed during abdominal insufflation when compared between different IAPs and between the two groups. Overall, an increased IAP of up to 20 mmHg did not significantly affect cardiovascular and respiratory variables in both the control and dexmedetomidine groups. This study suggests that the administration of a dexmedetomidine infusion is applicable in laparoscopic procedures in healthy dogs.

Clinical features and management of snake bites in 70 dogs in Korea.

BACKGROUND: Snakebites remain a devastating and life-threatening environmental hazard. While the management of snakebites has been well described in humans, few clinical data and guidelines exist for dogs, especially in Korea. OBJECTIVES: This retrospective study evaluated the clinical features of 70 dogs with snakebite wounds in Korea. METHODS: The medical records of 72 dogs that presented to three animal hospitals from June 2008 to July 2021 were reviewed; among these, 70 dogs that met the inclusion criteria were enrolled. Their signalment, history, clinical signs, physical examination, blood analysis, treatment, and prognosis were also evaluated. RESULTS: Of 70 dog owners, 35 (50%) witnessed the bite, with a mean time between bite and hospital presentation of 9.7 ± 4.1 h in 58 dogs. Blood smears were evaluated in 45 dogs, of which 28 (62%) showed echinocytosis. Anemia and acute kidney injury were found in 21 (29%) and 2 dogs (3%), respectively. A total of 37 dogs (53%) were hospitalized, 5 (7%) of which died. CONCLUSIONS: The most significant finding was the high prevalence of echinocytosis. The data from this retrospective study could inform the management of dogs bitten by snakes in Korea.

Lemon Balm and Corn Silk Mixture Alleviates Metabolic Disorders Caused by a High-Fat Diet.

We recently reported that varying combination ratios of lemon balm (Mellissa officinalis L.) and corn silk extracts (Stigma of Zea mays L. fruit) could reduce the obesity caused by a high-fat diet (HFD). The present study investigated the dose-dependent effect of a 1:1 (w:w) mixture of lemon balm and corn silk extracts (M-LB/CS) on HFD-mediated metabolic disorders and compared the effect with metformin. Oral administration of 50-200 mg/kg of M-LB/CS for 84 days significantly inhibited HFD-induced body weight gain, adipocyte hypertrophy, and lipogenic gene induction without affecting food consumption in mice. Biochemical analyses showed that M-LB/CS blocked abnormal lipid accumulation in the blood by escalating fecal lipid excretion. In addition, M-LB/CS prevented HFD-mediated pancreatic atrophy, decreased the number of insulin- and glucagon-immunoreactive cells, and inhibited increases in glycated hemoglobin, glucose, and insulin. Moreover, M-LB/CS also reduced hepatic injury, lipid accumulation, gluconeogenesis, and lipid peroxidation in parallel with the induction of AMP-activated protein kinase and antioxidant enzymes. Furthermore, M-LB/CS protected the kidney by inhibiting tubular vacuolation and reducing serum creatinine and blood urea nitrogen levels. The prophylactic effect of 100 mg/kg M-LB/CS-administration was comparable to that of metformin. Therefore, M-LB/CS may be an alternative option for managing obesity and its related metabolic disorders.

Isolation and Characterization of Feline Wharton's Jelly-Derived Mesenchymal Stem Cells.

Wharton's jelly is a well-known mesenchymal stem cell source in many species, including humans. However, there have been no reports confirming the presence of mesenchymal stem cells in Wharton's jelly in cats. The purpose of this study was to isolate mesenchymal stem cells (MSCs) from the Wharton's jelly of cats and to characterize stem cells. In this study, feline Wharton's jelly-derived mesenchymal stem cells (fWJ-MSCs) were isolated and successfully cultured. fWJ-MSCs were maintained and the proliferative potential was measured by cumulative population doubling level (CPDL) test, scratch test, and colony forming unit (CFU) test. Stem cell marker, karyotyping and immunophenotyping analysis by flow cytometry showed that fWJ-MSCs possessed characteristic mesenchymal stem cell markers. To confirm the differentiation potential, we performed osteogenic, adipogenic and chondrogenic induction under each differentiation condition. fWJ-MSCs has the ability to differentiate into multiple lineages, including osteogenic, adipogenic and chondrogenic differentiation. This study shows that Wharton's jelly of cat can be a good source of mesenchymal stem cells. In addition, fWJ-MSCs may be useful for stem cell-based therapeutic applications in feline medicine.

Lemon Balm and Corn Silk Extracts Mitigate High-Fat Diet-Induced Obesity in Mice.

Lemon balm and corn silk are valuable medicinal herbs, which exhibit variety of beneficial effects for human health. The present study explored the anti-obesity effects of a mixture of lemon balm and corn silk extracts (M-LB/CS) by comparison with the effects of single herbal extracts in high-fat diet (HFD)-induced obesity in mice. HFD supplementation for 84 days increased the body weight, the fat mass density, the mean diameter of adipocytes, and the thickness of fat pads. However, oral administration of M-LB/CS significantly alleviated the HFD-mediated weight gain and adipocyte hypertrophy without affecting food consumption. Of the various combination ratios of M-LB/CS tested, the magnitude of the decreases in weight gain and adipocyte hypertrophy by administration of 1:1, 1:2, 2:1, and 4:1 (w/w) M-LB/CS was more potent than that by single herbal extracts alone. In addition, M-LB/CS reduced the HFD-mediated increases in serum cholesterol, triglyceride, and low-density lipoprotein, prevented the reduction in serum high-density lipoprotein, and facilitated fecal excretion of cholesterol and triglyceride. Moreover, M-LB/CS mitigated the abnormal changes in specific mRNAs associated with lipogenesis and lipolysis in the adipose tissue. Furthermore, M-LB/CS reduced lipid peroxidation by inhibiting the HFD-mediated reduction in glutathione, catalase, and superoxide dismutase. Therefore, M-LB/CS is a promising herbal mixture for preventing obesity.

Mesenchymal Stem Cell Exosomes Derived from Feline Adipose Tissue Enhance the Effects of Anti-Inflammation Compared to Fibroblasts-Derived Exosomes.

Adipose tissue-derived mesenchymal stem cells (AD-MSCs) release extracellular vesicles such as exosomes, apoptotic bodies, and microparticles. In particular, exosomes are formed inside cells via multivesicular bodies (MVBs), thus their protein, DNA, and RNA content are similar to those of the parent cells. Exosome research is rapidly expanding, with an increase in the number of related publications observed in recent years; therefore, the function and application of MSC-derived exosomes could emerge as cell-free therapeutics. Exosomes have been isolated from feline AD-MSCs and feline fibroblast cell culture media using ultracentrifugation. Feline exosomes have been characterized by FACS, nanoparticle tracking analysis, and transmission electron microscopy imaging. Moreover, cytokine levels were detected by sandwich enzyme-linked immunosorbent assay in exosomes and LPS-induced THP-1 macrophages. The size of the isolated exosomes was that of a typical exosome, i.e., approximately 150 nm, and they expressed tetraspanins CD9 and CD81. The anti-inflammatory factor IL-10 was increased in feline AD-MSC-derived exosomes. However, pro-inflammatory factors such as IL-1ß, IL-8, IL-2, RANTES, and IFN-gamma were significantly decreased in feline AD-MSC-derived exosomes. This was the first demonstration that feline AD-MSC-derived exosomes enhance the inflammatory suppressive effects and have potential for the treatment of immune diseases or as an inflammation-inhibition therapy.

The change of cytokines in tear and blood after different pterygium operation.

Pterygium is an invasion of altered ocular tissue into the cornea. Bone marrow-derived stem cells have been reported to be involved in wound healing under chemotactic factors after pterygium removal and pain may act as a trigger signal. We evaluated the change of systemic and local chemotactic factors that could affect the mobilization and migration of BMSCs to the wound bed after conventional bare sclera pterygium excision. We also applied temporary amniotic membrane patch after pterygium removal, and compared the changes of cytokines with those of conventional bare sclera excision group. Substance-P (SP), vascular endothelial growth factor (VEGF), and stem cell factor (SCF) were measured in plasma and tear using ELISA and migrating CD34(+) cells by flow cytometry. The results showed that post-operative pain was much reduced (p<0.05), and SP, VEGF and SCF kept consistently lower levels in plasma after temporary amniotic membrane application. Circulating CD34(+) cells increased slightly in the temporary amniotic membrane patch group compared with marked increase in the bare sclera group. Thus, the application of a temporary amniotic membrane after pterygium removal might be an effective therapeutic means by controlling pain and excessive infiltration of bone marrow-derived stem cells.

Cementless total hip arthroplasty performed in patients with pigmented villonodular synovitis.

The authors retrospectively reviewed all patients with pigmented villonodular synovitis (PVNS) of the hip treated by cementless total hip arthroplasty (THA) combined with synovectomy. Eight patients were reviewed and observed for an average of 8.9 years (range, 4.3-13.5 years). Mean preoperative Harris hip scores improved from 49.3 to 96.6 points, and all patients were able to regularly perform moderate daily living activities. None of the patients had clinical or radiographic evidence of recurrent PVNS. Osteolysis occurred in 4 hips, and 2 revision surgeries were performed during follow-up. Cementless THA combined with synovectomy is an adequate therapeutic choice for patients with PVNS demonstrating end-stage joint destruction and appears to be effective at improving clinical results and preventing disease recurrence.

Immunomodulatory effects of Aureobasidium pullulans SM-2001 exopolymers on the cyclophosphamide-treated mice.

The immunomodulatory effects of exopolymers of Aureobasidium pullulans SM-2001 containing beta-1,3/1,6-glucan were evaluated on the cyclophosphamide (CPA)-treated mice. To induce immunosuppress, 150 and 110 mg/kg of CPA were intraperitoneally injected at 1 and 3 days before start of test material administrations, respectively. Exopolymers were subcutaneously or orally administered in a volume of 10 ml/kg, 4 times; 12-hr intervals from 24 hrs after second treatment of CPA. After treatment of exopolymers, the changes of thymus and spleen weights, splenic amounts of tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta and IL-10, thymic and splenic CD3+, CD4+, CD8+ and TNF-alpha+ cells were monitored in CPA-treated mice. As results of CPA treatment, dramatical decreases of the CD3+, CD4+, CD8+ and TNF-alpha+ cells were detected in thymus and spleen with decreases of thymus and spleen weights. In addition, decreases of splenic TNF-alpha, IL-1beta and IL-10 contents were also detected at flow cytometrical observations. However, oral and subcutaneous treatment of exopolymers effectively reduced the immunosuppressive changes induced by CPA. Therefore, it is concluded that exopolymers of A. pullulans can be effectively prevent the immunosuppress mediated, at least partially, recruitment of T cells and TNF-alpha+ cells or enhancement of their activity, and can provide effective prevention or treat regimes for the immunosuppress and related diseases such as cancer, sepsis and high-dose chemotherapy or radiotherapy.

Effects of a herbal formulation, KGC3P, and its individual component, nepetin, on coal fly dust-induced airway inflammation.

Coal fly dust (CFD)-induced asthma model is used as an ambient particulate matter model of serious pulmonary damage. We aimed to evaluate the effects of a combination of ginseng and Salvia plebeia R. Br extract (KGC-03-PS; KG3P) and its individual components (hispidulin, nepetin and rosmarinic acid) in a CFD-induced mouse model of airway inflammation (asthma). We also evaluated signal transduction by KG3P and its individual components in the alveolar macrophage cell line, MH-S cells. In vitro, KG3P and its individual components inhibited nitric oxide production and expression of pro-inflammatory mediators and cytokines (iNOS, COX-2, IL-1ß, IL-6 and TNF-α) through the NF-κB and MAPK pathways in coal fly ash (CFA)-induced inflammation in MH-S cells. Moreover, in the CFD-induced asthma model in mice, KG3P and its predominant individual component, nepetin, inhibited Asymmetric Dimethyl arginine (ADMA) and Symmetric Dimethyl arginine (SDMA) in serum, and decreased the histopathologic score in the lungs. A significant reduction in the neutrophils and immune cells in BALF and lung tissue was demonstrated, with significant reduction in the expression of the pro-inflammatory cytokines. Finally, IRAK-1 localization was also potently inhibited by KG3P and nepetin. Thus, KG3P extract can be considered as a potent candidate for amelioration of airway inflammation.

Ribonuclease inhibitor regulates neovascularization by human angiogenin.

Human angiogenin (ANG) is a homologue of bovine pancreatic ribonuclease (RNase A) that induces neovascularization. ANG is the only human angiogenic factor that possesses ribonucleolytic activity. To stimulate blood vessel growth, ANG must be transported to the nucleus and must retain its catalytic activity. Like other mammalian homologues of RNase A, ANG forms a femtomolar complex with the cytosolic ribonuclease inhibitor protein (RI). To determine whether RI affects ANG-induced angiogenesis, we created G85R/G86R ANG, which possesses 10(6)-fold lower affinity for RI but retains wild-type ribonucleolytic activity. The neovascularization of rabbit corneas by G85R/G86R ANG was more pronounced and more rapid than by wild-type ANG. These findings provide the first direct evidence that RI serves to regulate the biological activity of ANG in vivo.

One-stage cementless revision arthroplasty for infected hip replacements.

We hypothesised that one-stage cementless revision hip arthroplasty may have advantages and a role in the treatment of selected patients with an infected hip replacement. We retrospectively reviewed all patients with an infected hip replacement treated with one-stage revision using cementless implants. Twelve patients were reviewed and followed up for at least three years (average: 7.2 years, range: 3.3-11.3 years) postoperatively. One recurrence of infection and one aseptic stem loosening were detected during follow-up. Grafted bone, which was used in eight patients, appeared to have united to host bone in all patients. The success rate of treatment was 83.3% when infection recurrence or component loosening were regarded as failure. One-stage revision hip arthroplasty using cementless implants appears to have a role in the treatment of carefully selected patients with an infected hip replacement if meticulous débridement is performed and appropriate antibiotics are properly used.

Prevention of cold ischemia/rewarming-induced ERK 1/2, p38 kinase and HO-1 activation by trophic factor supplementation of UW solution.

We have previously shown that trophic factor supplementation (TFS) of University of Wisconsin (UW) solution reduced early apoptotic changes in vascular endothelial cells. Here, we examine the effect of TFS on cell signaling pathways related to cell growth, differentiation, and apoptosis after cold ischemic storage. In this study, the effect of TFS on the phosphorylation of signaling molecules ERK (extracellular regulated-signaling kinase) 1/2 and p38 MAPK (mitogen activated protein kinases) and of HO-1 (hemeoxygenase-1), relative to changes seen in unmodified UW solution, were determined by Western blot in cells stored under cold ischemic conditions. Primary cultures of canine kidney proximal tubule cells (CKPTC) and human umbilical vein endothelial cells (HUVEC) were used in this study. There was a significant decrease, relative to UW solution, after 1 min rewarming in ERK 1 and 2 activity in CKPTCs. For p38 MAPK, a significant decrease after 5 min rewarming was seen in CKPTC (p<0.05) while significant reductions relative to UW solution were seen in HUVECs after both 1 and 5 min rewarming (p<0.05). Phosphorylated HO-1 was also decreased by 43% and 50% in HUVECs, relative to UW solution, after 1 and 5 min rewarming (p<0.05 at each time point). Collectively, TFS not only limits ERK 1/2 and p38 MAPK activity induced by cold ischemic injury and subsequent rewarming, but also substantially restricted increases in HO-1 phosphorylation.

The effect of trophic factor supplementation on cold ischemia-induced early apoptotic changes.

We have previously shown that trophic factor supplementation (TFS) of University of Wisconsin (UW) solution enhanced kidney viability after cold storage. Here, we use an in vitro model to study the effect of TFS on early apoptotic changes after cold ischemic storage. Mitochondrial membrane potential was determined by fluorescence intensity in primary canine kidney tubule cells, Madin-Darby canine kidney cells, and human umbilical vein endothelial cells. In addition, caspase 3 enzyme activity assay and immunofluorescence staining were performed to evaluate apoptosis. There was a 15% increase in mitochondrial membrane potential in human umbilical vein endothelial cells stored in trophic factor supplemented University of Wisconsin solution after four-hour rewarming (P<0.05). TFS suppressed caspase 3 enzyme activity and activation in human umbilical vein endothelial cells. We confirmed that the presence of TFS in UW solution has a beneficial effect by protecting mitochondrial function and reducing early apoptotic changes in vascular endothelial cells.