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.

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.

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.

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.

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.

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.

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.

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.

Biodegradation and Biocompatibility of Poly L-lactic Acid Implantable Mesh.

PURPOSE: The purpose of this study was to evaluate the biodegradation and biocompatibility of poly L-lactic acid (PLLA) implantable mesh under in vitro and in vivo conditions. METHODS: PLLA mesh was examined for changes in weight and tensile strength. The histology of the tissue around the PLLA implant was also evaluated. RESULTS: The weight and tensile strength of the PLLA prosthesis was stable for 180 days. In addition, the surface of the PLLA mesh was not digested under in vitro or in vivo conditions as determined by scanning electron microscope. Histologically, there were no significant changes in the diameters of implanted PLLA mesh and subtype fibers over the course of 180 days. Likewise, there were no significant changes in the number of inflammatory and mast cells after 180 days, nor was there an increase in the percentage of collagen surrounding the PLLA mesh. CONCLUSIONS: The results indicate that PLLA prostheses have good rigidity and biocompatibility in vivo and in vitro.

Congenital lobar emphysema concurrent with pneumothorax and pneumomediastinum in a dog.

A two-year-old castrated male Pomeranian dog was referred with the chief complaints of coughing and subcutaneous emphysema. On physical examination, the crepitant areas were palpable. When auscultated, the right chest was absent of respiratory sound, while the sound of the opposite side was enhanced. Radiographs presented pneumothorax and pneumomediastinum. On computed tomography, hypoattenuated bulla-like lesion at right middle lung lobe and trapped air in mediastinum were shown. After patient stabilization, surgery for excision of affected lobe was performed. During follow-up period, there were no recurrence and complication on radiographic examination. Based on clinical and pathological findings, the dog was diagnosed as congenital lobar emphysema.

Adipose-derived mesenchymal stem cells and platelet-rich plasma synergistically ameliorate the surgical-induced osteoarthritis in Beagle dogs.

BACKGROUND: The purpose of this study is to investigate the clinical effects of platelet-rich plasma (PRP) and adipose-derived mesenchymal stem cell (MSC) as the fundamental treatment of osteoarthritis (OA). METHODS: Twenty four Beagle dogs were used as cranial cruciate ligament transection models. The dogs were divided into four groups (n = 6) according to the intra-articular injection materials: the control group with phosphate-buffered saline (PBS), the PRP group with PRP, the MSC group with MSCs emerged in PBS, and the MSC and PRP co-treatment (MP) group with MSCs and PRP. RESULTS: Lameness score, focal compression strength, articular extracellular matrix (ECM) compositions, histopathology, and real-time PCR were used to evaluate the effects of PRP and MSCs on canine OA. In the order of MP, PRP, and MSC group, these all showed positive effects on the evaluated categories. The lameness scores were lower, and the focal compression strengths of the affected femoral articular surface cartilages were higher than those in the OA control group. Also, the inflammatory changes, when evaluated with Mankin scoring and histomorphologic examination, were significantly ameliorated with the treatment of PRP and/or MSCs. The glycosaminoglycan and collagen composition of extracellular matrix was more favorable in the test groups. The ECM-related genes significantly increased through the up-regulation, while the protein expressions of inflammatory cytokines were decreased through the inhibitory effects of PRP and MSCs on chondrocyte apoptosis and inflammatory cytokines. CONCLUSIONS: Taken together, this study suggests that PRP and MSCs treatments have a beneficial effect on OA via the stimulation of ECM synthesis and chondrocyte proliferation and via the inhibition of inflammatory reaction.

Therapeutic effect of irradiation of magnetic infrared laser on osteoarthritis rat model.

Osteoarthritis (OA) is a degenerative joint disease caused by articular cartilage loss. Many complementary and alternative medicines for OA have been reported so far, but the effectiveness is controversial. Previously, we have shown anti-inflammatory effects of low level laser therapy with static magnetic field, magnetic infrared laser (MIL), in various animal models. Therefore, the beneficial effects were examined in OA rat model. Rats were divided by six groups; no treatment controls of sham and OA model, three MIL treatment groups of OA model at 6.65, 2.66 and 1.33 J cm(-2), and Diclofenac group of OA model with 2 mg kg(-1) diclofenac sodium. The OA control exhibited typical symptoms of OA, but 4-week MIL treatment improved the functional movement of knee joint with reduced edematous changes. In addition, cartilage GAGs were detected more in all MIL treatment groups than OA control. It suggests that 4-week MIL irradiation has dose-dependent anti-inflammatory and chondroprotective effects on OA. Histopathological analyses revealed that MIL treatment inhibits the cartilage degradation and enhances chondrocyte proliferation. The fact that MIL has an additional potential for the cartilage formation and no adverse effects can be regarded as great advantages for OA treatment. These suggest that MIL can be useful for OA treatment.

Nephroprotective Effect of POLYCAN on Acute Renal Failure Induced by Cisplatin in Rats.

We performed to evaluate the effect of POLYCAN (ß-glucan) on cisplatin-(CDDP-)induced acute renal failure (ARF) in rats. POLYCAN was administered orally once a day for 32 days. Each of 8 rats per group was selected based on the body weight (BW) after acclimatization and they were sacrificed at 5 days after CDDP injection. There was significant (P < 0.05) increase of BW after CDDP dosing in all POLYCAN groups than vehicle control and significant (P < 0.01 or P < 0.05) decrease of absolute and relative kidney weight were detected in all POLYCAN groups compared with vehicle control. In addition, serum BUN and creatinine level in all POLYCAN groups were significantly (P < 0.01 or P < 0.05) lower than vehicle control and the percentage of degenerative regions significantly (P < 0.01) decreased in all POLYCAN groups. As the results of CDDP-induced ARF process, dramatic decrease of the BW, increase of the kidney weight, serum BUN, and creatinine level were detected in vehicle control group compared with sham control group. The changes by CDDP-induced ARF process in POLYCAN groups were significantly and dose-dependently improved compared with vehicle control group. Therefore, POLYCAN has enough potential to develop as a new agent of prevention or treatment for ARF.

Fermented garlic protects diabetic, obese mice when fed a high-fat diet by antioxidant effects.

This study examined the bioactivity of yeast (Saccharomyces cerevisiae)-fermented aged black garlic (FBG) on obese mice supplied a high-fat diet (HFD) and its in vitro antioxidant activity. Aged black garlic (BG) exhibits potent antioxidative effects and has been subjected to extensive research. In addition, the bioactivity of some natural products is increased by fermentation. In a preliminary test, this study found that the antioxidant activity of FBG is stronger than that of BG. Therefore, it was hypothesized that the bioactivity of BG would be increased by yeast fermentation and would be a good candidate as a nutraceutical product for improving the oxidative defense systems in older patients or patients affected by various oxidative stresses, for example, diabetes and diabetic complications. To test this hypothesis, the bioactivities of FBG in diabetic and obese mice as well as the antioxidant activity in vitro were examined. After 91 days of continuous HFD supply, the mice showed marked obesity, hyperglycemia, hyperlipemia, and liver and kidney damages. Black garlic and all 3 different doses of FBG showed favorable hepatoprotective, nephroprotective, hypolipidemic, and antiobesity effects compared with the HFD control, but no hypoglycemic effects. In particular, more favorable bioactivity against all 4 HFD-induced diabetic complications was detected in the FBG-treated groups compared with the group given equivalent doses of BG. These findings suggest that the bioactivities of BG can be improved by yeast fermentation.

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.

Transplantation of reconstructed corneal layer composed of corneal epithelium and fibroblasts on a lyophilized amniotic membrane to severely alkali-burned cornea.

The purpose of this article was to evaluate the graft efficacy of reconstructed corneal layer, composed of autologous corneal epithelium and fibroblasts on a lyophilized amniotic membrane (LAM), in a severely alkali-burned corneal model. After biopsy specimens were obtained from the left eyes of 24 rabbits, the corneal epithelial cells and fibroblasts were expanded in vitro and the corneal layer was reconstructed on LAM. Thirty-six eyes of rabbits underwent alkali burn (1 N NaOH, 30 s) to create a limbal deficiency and a deeply damaged corneal stroma. Four weeks later, group 1 underwent a graft of the reconstructed corneal layer composed of autologous corneal epithelium and fibroblasts on LAM. Group 2 was transplanted with a graft of the reconstructed autologous corneal epithelium, and group 3 served as a control without surgery. Wound healing and stabilization of the ocular surfaces occurred much faster in group 1 than in groups 2 and 3. The eyes in group 3 revealed typical limbal deficiencies with conjuctivalization and persistent corneal epithelial defects. However, the corneas in group 1 developed only mild peripheral neovascularization. Immunohistochemical staining in group 1 demonstrated that p63, cytokeratin 3, E-cadherin, transforming growth factor (TGF)-beta1, and collagen IV were expressed strongly in the corneal epithelium and basement membrane. On the basis of these results, transplantation of the reconstructed corneal layer, composed of autologous corneal epithelium and fibroblasts on LAM, partially accelerated the recovery of the alkali-injured rabbit ocular surface, and might be useful therapeutically for the treatment of patients with severely damaged cornea.