Mesenchymal Stem Cells and Extracellular Vesicles Derived from Canine Adipose Tissue Ameliorates Inflammation, Skin Barrier Function and Pruritus by Reducing JAK/STAT Signaling in Atopic Dermatitis.

Canine atopic dermatitis (AD) is a common chronic inflammatory skin disorder resulting from imbalance between T lymphocytes. Current canine AD treatments use immunomodulatory drugs, but some of the dogs have limitations that do not respond to standard treatment, or relapse after a period of time. Thus, the purpose of this study was to evaluate the immunomodulatory effect of mesenchymal stem cells derived from canine adipose tissue (cASCs) and cASCs-derived extracellular vesicles (cASC-EVs) on AD. First, we isolated and characterized cASCs and cASCs-EVs to use for the improvement of canine atopic dermatitis. Here, we investigated the effect of cASCs or cASC-EVs on DNCB-induced AD in mice, before using for canine AD. Interestingly, we found that cASCs and cASC-EVs improved AD-like dermatitis, and markedly decreased levels of serum IgE, (49.6%, p = 0.002 and 32.1%, p = 0.016 respectively) epidermal inflammatory cytokines and chemokines, such as IL-4 (32%, p = 0.197 and 44%, p = 0.094 respectively), IL-13 (47.4%, p = 0.163, and 50.0%, p = 0.039 respectively), IL-31 (64.3%, p = 0.030 and 76.2%, p = 0.016 respectively), RANTES (66.7%, p = 0.002 and 55.6%, p = 0.007) and TARC (64%, p = 0.016 and 86%, p = 0.010 respectively). In addition, cASCs or cASC-EVs promoted skin barrier repair by restoring transepidermal water loss, enhancing stratum corneum hydration and upregulating the expression levels of epidermal differentiation proteins. Moreover, cASCs or cASC-EVs reduced IL-31/TRPA1-mediated pruritus and activation of JAK/STAT signaling pathway. Taken together, these results suggest the potential of cASCs or cASC-EVs for the treatment of chronic inflammation and damaged skin barrier in AD or canine AD.

Antioxidative and antiproliferative activities of ethanol extracts from pigmented giant embryo rice (Oryza sativa L. cv. Keunnunjami) before and after germination.

BACKGROUND/OBJECTIVES: Oxidative stress is a major cause of cancer. This study investigated the effects of the ethanol extracts from germinated and non-germinated Keunnunjami rice, a blackish-purple pigmented cultivar with giant embryo, on selected human cancer cell lines and on the antioxidant defense system of mice fed with a high-fat diet. MATERIALS/METHODS: High fat-fed mice were orally administered with either distilled water (HF) or extracts (0.25%, w/w) from brown (B), germinated brown (GB), Keunnunjami (K), and germinated Keunnunjami (GK) rice. RESULTS: In comparison with the brown rice extract, Keunnunjami extract showed higher anticancer effect against cervical and gastric cell lines but lower anticancer activity on liver and colon cancer cells. Mice from the HF group showed significantly higher lipid peroxidation and lower antioxidant enzyme activities than the control group. However, the oxidative stress induced by high-fat diet markedly decreased in B, GB, K, and GK groups as compared with the HF group. CONCLUSIONS: Germination may be an effective method for improving the anticancer and antioxidative properties of Keunnunjami rice and extracts from germinated Keunnunjami rice may serve as a therapeutic agent against cervical and gastric cancers and oxidative damage.

Dietary supplementation of germinated pigmented rice (Oryza sativa L.) lowers dyslipidemia risk in ovariectomized Sprague-Dawley rats.

BACKGROUND: In the recent years, cases of elderly women suffering from metabolic diseases such as dyslipidemias brought about by hormonal imbalance after menopause are continuously increasing. In this regard, a continuous and escalating demand to develop a more functional and highly nutritional food product as an adjunct supplement that can help alleviate these diseases is still being sought. OBJECTIVE: This study investigated the effects of germinated blackish-purple rice cultivars Keunnunjami, Superjami, and reddish-brown cultivar Superhongmi in the lipid metabolism of ovariectomized Sprague-Dawley rats. METHOD: The animals were randomly divided into nine groups (n=5) and were supplemented with either non-germinated or germinated rice for 9 weeks. Then the plasma, liver, and fat samples were collected for the lipid metabolism effects analyses. RESULTS: Animals fed with germinated rice cultivars had improved lipid profile levels relative to the groups supplemented with non-germinated rice cultivars. The germinated rice groups, Keununjami and Superjami in particular, showed a low total cholesterol levels, high levels of high-density lipoproteins-cholesterol, high fecal lipid output, low hepatic lipid values, and low hepatic adipocyte accumulation. There was also an increase in the rate of lipolysis and decrease in lipogenesis based on the lipid-regulating enzyme activity profiles obtained for the groups that fed on germinated rice. Also, results revealed that pigmented rice cultivars had superior effects in improving the lipid metabolism relative to the non-pigmented normal brown rice variety. CONCLUSION: Based on the results, this study suggests that germinated pigmented rice consumption can confer better lipid metabolism than ordinary white rice and constitutes as an effective functional food in alleviating the risk of having dyslipidemias like those suffering from menopausal co-morbidities.

Antihyperlipidemic effects of Korean ginseng in high-fat diet-fed ovariectomized rats.

Because of decreased estrogen levels, postmenopausal women are at a high risk of developing dyslipidemia, obesity, and other metabolic diseases. The effects of Korean ginseng were investigated to determine whether it can be used as an adjunct supplement for hyperlipidemia. Ovariectomized Sprague-Dawley rats were used to mimic postmenopausal conditions and were randomly divided into five groups (n=6): normal control, high-fat diet, high-fat diet+statin, high-fat diet+white ginseng, and high-fat diet + heated ginseng. In vitro results revealed that heated ginseng had higher ginsenosides. Meanwhile, in vivo results showed that heated ginseng inhibited excessive lipogenesis and promoted lipolysis. It also had a better lipid profile than white ginseng. Finally, plasma adipokine levels indicated a lower risk of developing obesity-induced hyperlipidemia in rats fed heated ginseng. Overall, these results suggest that heated ginseng ameliorates hyperlipidemia and that it can be a suitable alternative to white ginseng.

Red-koji fermented red ginseng ameliorates high fat diet-induced metabolic disorders in mice.

Fermentation of medicinal herbs improves their pharmacological efficacy. In this study, we investigated the effects of red-koji fermented red ginseng (fRG) on high-fat diet (HFD)-mediated metabolic disorders, and those effects were compared to those of non-fermented red ginseng (RG). fRG (500, 250 or 125 mg/kg), RG (250 mg/kg), simvastatin (10 mg/kg), silymarin (100 mg/kg) and metformin (250 mg/kg) were orally administered from 1 week after initiation of HFD supply for 84 days. The diameter of adipocytes in periovarian and abdominal fat pads and the thickness of the abdominal fat were significantly decreased by fRG treatment, while HFD-mediated weight gain was partly alleviated by fRG in a dose-dependent manner. Moreover, biochemical and histomorphometrical analyses clearly indicated that fRG significantly inhibited HFD-induced metabolic disorders such as hyperglycemia, hyperlipidemia, hepatopathy and nephropathy in a dose-dependent manner. More favorable pharmacological effects on HFD-mediated metabolic disorders were also observed with fRG compared to an equal dose of RG. This finding provides direct evidence that the pharmacological activities of RG were enhanced by red-koji fermentation, and fRG could be a neutraceutical resource for the alleviation of obesity-mediated metabolic disorders.

Tamoxifen-induced activation of p21Waf1/Cip1 gene transcription is mediated by Early Growth Response-1 protein through the JNK and p38 MAP kinase/Elk-1 cascades in MDA-MB-361 breast carcinoma cells.

Tamoxifen (TAM) is a synthetic non-steroidal anti-estrogen compound that is widely used as an effective chemotherapeutic agent for treatment and prevention of breast cancer. Unfortunately, prolonged treatment with TAM causes TAM-responsive tumors to become TAM resistant through an as-yet-unknown mechanism. To develop novel anti-breast cancer agents that are therapeutically superior to TAM, we must first fully understand the biological effects of TAM. In this study, we found that TAM treatment of MDA-MB-361 breast cancer cells activated p21Waf1/Cip1 gene transcription independently of p53. Furthermore, TAM-induced p21Waf1/Cip1 promoter activity was enhanced by transient expression of the gene encoding Early Growth Response-1 (Egr-1) protein, a transcription factor that plays an important role in cell growth and differentiation. The TAM-induced p21Waf1/Cip1 promoter activity was blocked by the expression of small interfering RNA (siRNA) targeted to Egr-1 mRNA. In addition, induction of Egr-1 expression by TAM occurred at the transcriptional level via Ets-domain transcription factor Elk-1 through the JNK and p38 mitogen-activated protein (MAP) kinase pathways. Inhibition of the JNK and p38 MAP kinase signals inhibited Egr-1-mediated p21Waf1/Cip1 promoter activity. We conclude that TAM stimulation of p21Waf1/Cip1 gene transcription in MDA-MB-361 cells depends largely on Elk-1-mediated Egr-1 expression induced by activation of the JNK and p38 MAP kinase pathways.

Changes in orexin-A and neuropeptide y expression in the hypothalamus of obese and lean Zucker Diabetic Fatty rats.

This study was carried out to investigate the changes of orexin-A (OXA) and neuropeptide Y (NPY) expression in the hypothalamus of the obese and lean Zucker Diabetic Fatty (ZDF) rats which have a missense mutation in the leptin receptor gene. The mean body weights (MBW) between the obese and lean ZDF rats were significantly different at 28 and 70 postnatal days. However, at 14 postnatal day, there was no significant difference in the MBW between the obese and lean ZDF rats in both male and female. The OXA immunoreactivities were not significantly different between the obese and lean ZDF rats in both sexes at 14, 28, and 70 postnatal days, respectively. The NPY immunoreactivity was higher in the obese than in the lean ZDF rats in both male and female at 28 and 70 postnatal days, whereas there was no significant difference between the obese and lean ZDF rats at 14 postnatal day. These results indicate that both OXA and NPY might halt their roles for food intake in the obese phenotype of the male and female ZDF rats in the preweaning period of 14 postnatal day, whereas NPY might play a main role in the obesity of these rats in the weaning period of 28 and 70 postnatal days.

Neuroglial activation in Niemann-Pick Type C mice is suppressed by intracerebral transplantation of bone marrow-derived mesenchymal stem cells.

Glial activation is thought to play a key role in pathogenesis of neurodegenerative disorders. Here we show that direct transplantation of bone marrow-derived mesenchymal stem cells (BM-MSC) results in alleviation of inflammatory responses associated with the cerebellum of Niemann-Pick disease Type C (NP-C) model mice. Immunohistochemical examinations using glial fibrillary acidic protein (GFAP) and F4/80 antibodies revealed that BM-MSC transplantation reduced significantly both of astrocytic and microglial activations in the cerebellum of NP-C mice. Expression of macrophage colony stimulating factor (M-CSF), a microglial activator, was also considerably down-regulated by the BM-MSC transplantation. These findings suggest that BM-MSC transplantation may have potential for a therapeutic role in the treatment of NP-C and other neurodegenerative brain disorders.

Changes in orexin-A and neuropeptide Y expression in the hypothalamus of the fasted and high-fat diet fed rats.

This study was aimed to investigate the changes of orexin-A (OXA) and neuropeptide Y (NPY) expression in the hypothalamus of the fasted and high-fat diet fed rats. For the experiments, the male Sprague-Dawley (SD) rats were used as the model of high-fat diet-induced obesity. The mean loss of body weight (MLBW) did not show the linear pattern during the fasting; from 24 h to 84 h of fastings, the MLBW was not significantly changed. The numbers of OXA-immunoreactive (IR) neurons were decreased at 84 h of fasting compared with those in other five fasting subgroups. The NPY immunoreactivities in the arcuate nucleus (ARC) and the suprachiasmatic nucleus (SCN) observed at 84 h of fasting were higher than that observed at 24 h of fasting. The number of OXA-IR neurons of the LHA (lateral hypothalamic area) in the high-fat (HF) diet fed group was more increased than that of the same area in the normal-fat (NF) diet fed group. The NPY immunoreactivities of the ARC and the SCN were higher in HF group than those observed in the same areas of NF group. Based on these results, it is noteworthy that the decrease of the body weight during the fast was not proportionate to the time-course, implicating a possible adaptation of the body for survival against starvation. The HF diet might activate the OXA and the NPY in the LHA to enhance food intake.

Expressional changes of neuropeptide Y and cholecystokinin in the arcuate and paraventricular nuclei after capsaicin administration.

Despite its toxicity, a great deal of attention has been paid to the anorexic effect of capsaicin in the treatment of obesity-related neurotransmitters/neuromodulators. To determine if capsaicin has any effects on the orexigenic or anorexigenic peptides, the neuropeptide Y (NPY)- and cholecystokinin (CCK)-immunoreactivities were demonstrated in the rat hypothalamus by immunohistochemistry after capsaicin administration. There was a significantly lower concentration of NPY immunopositive cells in the arcuate and paraventricular nuclei of the capsaicin treated rats. In contrast, the CCK expressions level was higher in the paraventricular nucleus of the capsaicin treated rats than in the control rats. These results suggest that capsaicin influence neuropeptides such as orexigenic NPY and anorexigenic CCK related to control food intake.

Effect of capsaicin on cholecystokinin and neuropeptide Y expressions in the brain of high-fat diet fed rats.

Capsaicin, one of the pungent principles of hot pepper, has been reported to cause a cessation of increases in body weight and fat gain induced by high-fat feeding. Especially, in body weight and feeding control, cholecystokinin (CCK) has been well known as a satiety signal and neuropeptide Y (NPY) has been described as one of the most potent orexigenic signals. This study was carried out to investigate the effect of capsaicin on CCK- and NPY- immunoreactivities (IR) in the brain of high-fat fed rats. The animals were divided into normal-fat diet (NF), high-fat diet (HF) and high-fat diet containing capsaicin (HF-CAP) groups. Mean body weight gain (MBWG) of HF group was higher than that of NF group. However, in HF-CAP group, MBWG was lower than that of HF group. CCK-IR in suprachiasmatic nucleus (SCN), paraventricular nucleus (PVN), median eminence (ME), arcuate nucleus (ARC) and amygdala was not prominent in all the groups. In cerebral cortex, CCK-IR was more reduced in HF-CAP group than in the other groups. In the HF-CAP group, NPY-IR in the hypothalamic nuclei, amygdala and cerebral cortex was more poorly found than in the NF and HF groups. It is concluded that (1) NPY-IR may react more sensitively on capsaicin than CCK-IR, (2) no rapid increase of body weight in capsaicin treated rats may result from the diminished food intake through the low expression of NPY in hypothalamus in HF-CAP group.

Optimization of in situ hybridization protocols for detection of feline herpesvirus 1.

In situ hybridization (ISH) protocol including microwaving pre-treatment regimes was developed and compared with protease digestion as a pre-treatment regime for its effects on detecting feline herpesvirus 1 (FHV-1) in formalin fixed, paraffin embedded tissue. We found that optimum results were obtained using microwave pre-treatment. The results showed that the use of microwave irradiation would be recommended as a means of supplementing ISH methods, especially when using long-term formalin fixed, paraffin-embedded tissue.

GABA-, glycine-, and glutamate-immunoreactive bouton profiles in apposition to neurons of the central cervical nucleus in the rat.

The neurons of the central cervical nucleus (CCN) convey information about the position and movements of the head, and receive excitatory input from dorsal neck muscles and the labyrinth. Both of these afferent sources form glutamatergic synaptic contacts with CCN neurons. However, these sensory afferent sources can also inhibit CCN neurons. To further elucidate the synaptic organization, we made an electron microscopic investigation, identifying and evaluating the relative frequency of bouton profiles containing the inhibitory transmitters GABA and glycine in apposition to identified CCN neurons. In addition, labeling for glutamate was performed. The identification of the CCN neurons was made possible by injections of retrograde tracer substances into the cerebellum. These substances were made visible by preembedding immunocytochemistry or postembedding immunogold staining. Such staining was also used to detect the three amino acids that were found in boutons apposed to the identified neurons (cf. Ornung et al., J. Comp. Neurol. 1996;365:413-426; Lindå et al., J. Comp. Neurol. 2000;425:10-23). Due to the relatively poor transport of the tracer substances into dendrites of the CCN neurons, the analysis was restricted to the cell body and included bouton profiles in direct apposition to the soma membrane. Data from 10 CCN neurons revealed that about 50% of the apposing bouton profiles were immunoreactive for GABA, and about 34% for glycine. In four neurons, the degree of colocalization of GABA and glycine was determined to be close to 30%. Thus, the vast majority of glycine-labeled profiles also contained GABA, while a considerable fraction of the profiles were immunoreactive for only GABA. The values for glycine immunoreactive bouton profiles presented here may represent somewhat low estimates, depending on the method used. Data from four neurons showed that about 18% of the profiles were labeled for glutamate. The large fraction of purely GABA immunoreactive profiles, or at least a substantial group of them, is suggestive of their derivation from axons descending from the brainstem.