Omega-3 Fatty Acids Attenuate Renal Fibrosis via AMPK-Mediated Autophagy Flux Activation.

The unilateral ureteral obstruction (UUO) injury model is well-known to mimic human chronic kidney disease, promoting the rapid onset and development of kidney injury. ω3-poly unsaturated fatty acids (PUFAs) have been observed to protect against tissue injury in many disease models. In this study, we assessed the efficacy of ω3-PUFAs in attenuating UUO injury and investigated their mechanism of action. The immortalized human proximal tubular cells human kidney-2 (HK2) were incubated for 72 h with docosahexaenoic acid (DHA) or eicosapentaenoic acid (EPA) in various concentrations, in the presence or absence of transforming growth factor (TGF)-ß. DHA/EPA reduced the epithelial-mesenchymal transition in the TGF-ß-treated HK2 cells by enhancing autophagy flux and adenosine monophosphate-activated protein kinase (AMPK) phosphorylation. C57BL/6 mice were divided into four groups and treated as follows: sham (no treatment, n = 5), sham + ω3-PUFAs (n = 5), UUO (n = 10), and UUO + ω3-PUFAs (n = 10). Their kidneys and blood were harvested on the seventh day following UUO injury. The kidneys of the ω3-PUFAs-treated UUO mice showed less oxidative stress, inflammation, and fibrosis compared to those of the untreated UUO mice. Greater autophagic flux, higher amounts of microtubule-associated protein 1A/1B-light chain 3 (LC3)-II, Beclin-1, and Atg7, lower amounts of p62, and higher levels of cathepsin D and ATP6E were observed in the kidneys of the omega-3-treated UUO mice compared to those of the control UUO mice. In conclusion, ω3-PUFAs enhanced autophagic activation, leading to a renoprotective response against chronic kidney injury.

Ratio of Extracellular to Intracellular Water Is Associated with Permanent Catheter Patency Survival in Patients Receiving Maintenance Hemodialysis.

Patients undergoing dialysis through a permanent catheter often experience infection or malfunction. However, few studies have clarified the predictors of permanent catheter patency survival in patients undergoing hemodialysis. We assessed the relationship between the parameters of body composition monitoring (BCM), determined before the initiation of dialysis, and the patency survival of the permanent catheters inserted in 179 patients who commenced hemodialysis between 14 January 2020 and 31 August 2021. The relationships between permanent catheter patency at 6 weeks and BCM parameters, laboratory tests, age, sex, comorbidities, and medications at baseline were studied using Kaplan-Meier survival curves. Permanent catheter patency was observed to be superior at high extracellular-to-intracellular (ECW/ICW) ratio (p < 0.005). After adjustment for covariates, the ECW/ICW ratio remained an independent factor associated with permanent catheter patency survival. When patients with non-patent catheters were subdivided into infection and malfunction groups, and the associations of BCM parameters were evaluated in those groups, the ECW/ICW ratio was not significantly associated with permanent catheter patency survival in the infection group (p = 0.327); instead, a significant association was found for the lean tissue index (p < 0.001). In the malfunction group, the ECW/ICW ratio remained significantly associated with permanent catheter patency survival (p < 0.001).

Diagnostic and Prognostic Roles of C-Reactive Protein, Procalcitonin, and Presepsin in Acute Kidney Injury Patients Initiating Continuous Renal Replacement Therapy.

For reducing the high mortality rate of severe acute kidney injury (AKI) patients initiating continuous renal replacement therapy (CRRT), diagnosing sepsis and predicting prognosis are essential. However, with reduced renal function, biomarkers for diagnosing sepsis and predicting prognosis are unclear. This study aimed to assess whether C-reactive protein (CRP), procalcitonin, and presepsin could be used to diagnose sepsis and predict mortality in patients with impaired renal function initiating CRRT. This was a single-center, retrospective study involving 127 patients who initiated CRRT. Patients were divided into sepsis and non-sepsis groups according to the SEPSIS-3 criteria. Of the 127 patients, 90 were in the sepsis group and 37 were in the non-sepsis group. Cox regression analysis was performed to determine the association between the biomarkers (CRP, procalcitonin, and presepsin) and survival. CRP and procalcitonin were superior to presepsin for diagnosing sepsis. Presepsin was closely related to the estimated glomerular filtration rate (eGFR) (r = -0.251, p = 0.004). These biomarkers were also evaluated as prognostic markers. Procalcitonin levels ≥3 ng/mL and CRP levels ≥31 mg/L were associated with higher all-cause mortality using Kaplan-Meier curve analysis. (log-rank test p = 0.017 and p = 0.014, respectively). In addition, procalcitonin levels ≥3 ng/mL and CRP levels ≥31 mg/L were associated with higher mortality in univariate Cox proportional hazards model analysis. In conclusion, a higher lactic acid, sequential organ failure assessment score, eGFR, and a lower albumin level have prognostic value to predict mortality in patients with sepsis initiating CRRT. Moreover, among these biomarkers, procalcitonin and CRP are significant factors for predicting the survival of AKI patients with sepsis-initiating CRRT.

Artery-to-Fistula Diameter Ratio as a Predictor of Early Re-Occlusion of Immature Right Radio-Cephalic Arteriovenous Fistula after Primary PTA.

Percutaneous transluminal angioplasty (PTA) is widely performed for arteriovenous fistula (AVF) that fails to mature after initial formation. We observed that some immature AVFs re-occlude earlier than others. We sought to investigate the predictors for early post-intervention failure of immature fistulas after primary PTA. We retrospectively reviewed the records and angiographic images of patients who had immature fistulas and thereby received PTA between 2013 and 2019 at our center. We investigated the short-term post-intervention outcomes of the patients within 90 days post-PTA. Patients who had re-occlusion within the period were defined as the early failure group and the rest as the patent group. We investigated factors associated with early failure. There were 80 eligible patients with 22 brachio-cephalic (BC) and 58 radio-cephalic (RC) AVFs. The median age of the patients was 64 years [range, 38-87]. There were 51 (63%) males and 29 (36%) females. Among the 58 RC AVFs, 10 (17%) patients had early failure. Logistic regression analysis showed that a larger artery to fistula (A/F) diameter ratio was the sole independent predictor of early failure after primary PTA (odd ratio 2.29 [1.023-5.147], p value = 0.044). Although further studies on a larger scale are required to confirm the clinical significance, a larger A/F diameter ratio was a potential predictor of early re-occlusion in immature fistulas after primary PTA.

Serum Growth Differentiation Factor-15/Albumin Ratio as a 2-Year Survival Marker of End-Stage Renal Disease Patients Initiating Maintenance Hemodialysis.

It is important to identify risk factors related to mortality in end-stage renal disease (ESRD) patients starting renal replacement therapy. Recently, several studies proposed that growth-differentiation factor-15 (GDF-15) is a possible biomarker for the prognosis of patients on maintenance hemodialysis. Here, we investigated the predictive value of serum GDF-15/Albumin ratio on two-year mortality in ESRD patients initiating maintenance hemodialysis. The study was a single center, retrospective study on ESRD patients starting maintenance hemodialysis with a follow-up of two years. All patients completed laboratory test and bioimpedance spectroscopy prior to the initiation of the first dialysis. The patients were stratified into quartiles according to the quartiles of serum GDF-15/Albumin ratio. Among the 159 patients, the mean age was 61.78 ± 12.52 years and median survival was 20.03 ± 7.73 months. The highest GDF-15/Albumin quartile was significantly more associated with the increased risk of all-cause mortality than other quartiles (unadjusted hazard ratio (HR): 8.468, 95% CI 2.981-24.054, p < 0.001). Older age and a higher overhydration state were associated with GDF-15/Albumin ratio. The ROC analysis confirmed that the ability of the GDF-15/Albumin ratio to predict mortality was superior to GDF-15 or albumin alone. In conclusion, the GDF-15/Albumin ratio measured at the initial maintenance hemodialysis is an independent prognostic marker of two-year mortality in ESRD patients.

Progression to chronic kidney disease according to albumin uria in diabetic nephropathy patients with preserved renal function / Progresión a enfermedad renal crónica según albuminuria en pacientes con nefropatía diabética con función renal preservada

ABSTRACT Background : Albuminuria predicts progression of diabetic nephropathy (DN) but lacks specificity and sensitivity for the diagnosis of chronic kidney disease (CKD) and progressive decline in estimated glomerular filtration rate (eGFR). We evaluated the decline in renal function in patients with DN and analyzed the prognosis of renal function according to the level of albuminuria and the incidence of cardiovascular disease (CVD), cerebrovascular diseases, and peripheral artery disease (PAD) according to the level of albuminuria. Methods: This retrospective study included 331 patients with eGFR >60 mL/min/1.73 m2 and urinary albumin/creatinine (Cr) ratio (ACR) >30 mg/g Cr who were treated at the Chungnam National University Hospital between January 2012 and December 2018. Patients were divided into mildly increased albuminuria, moderately increased albuminuria, and severely increased albuminuria groups according to their urine ACRs of 30-300, 300 900, and >900 mg/g Cr, respectively. Renal outcomes and incidence of CVD, cerebrovascular disease, and PAD were compared among the three groups. Results: More severe albuminuria was associated with higher rates of progression to CKD (p< 0.001) and >50% reduction in eGFR from baseline (p< 0.001). There was a statistically significant difference in the rate of PCI with angina or myocardial infarction (p=0.030). However, cerebrovascular disease and PAD did not significantly differ among the three groups. Conclusión: Among patients with DN who maintained a relatively preserved renal function with an eGFR >60 mL/min/1.73 m2, the rates of renal deterioration and progression to CKD were significantly more frequent in those with more severe albuminuria.

RESUMEN Antecedentes: La albuminuria predice la progresión de la nefropatía diabética (ND) pero carece de especificidad y sensibilidad para el diagnóstico de la enfermedad renal crónica (ERC) y la disminución progresiva en la tasa de filtración glomerular estimada (eGFR). Evaluamos la disminución de la función renal en pacientes con ND y analizamos el pronóstico de la función renal de acuerdo con el nivel de albuminuria y la incidencia de enfermedad cardiovascular (ECV), enfermedades cerebrovasculares y enfermedad de las arterias periféricas (EAP) de acuerdo con el nivel de albuminuria. Material y métodos: Este estudio retrospectivo incluyó a 331 pacientes con eGFR >60 ml/min/1,73 m2 y de albúmina urinaria/creatinina (CR) (ACR) >30 mg/g CR que fueron tratados en el Hospital Universitario Nacional de Chungnam entre enero de 2012 y diciembre de 2018. Los pacientes se dividieron en tres grupos: albuminuria ligeramente aumentada, aumento moderado de albuminuria y aumento severo de albuminuria de acuerdo con sus ACRs de orina de 30-300, 300-900 y >900 mg/g Cr, respectivamente. Los resultados renales e incidencia de ECV, enfermedad cerebrovascular y EAP se compararon entre los tres grupos. Resultados: La albuminuria más severa se asoció con tasas más altas de progresión a ERC (P <0,001) y una reducción >50% en eGFR desde la línea de base (P <0,001). Hubo una diferencia estadísticamente significativa en la tasa de PCI con la angina o el infarto de miocardio (P =0,030). Sin embargo, la enfermedad cerebrovascular y la EAP no difirieron significativamente entre los tres grupos. Conclusión: entre los pacientes con ND que mantuvieron una función renal relativamente conservada con un eGFR >60 ml/ min/1,73 m2, las tasas de deterioro renal y la progresión a la ERC fueron significativamente más frecuentes en aquellos con albuminuria más severa.

Application of Gamma Ray-Responsive Genes for Transcriptome-Based Phytodosimetry in Rice.

Transcriptome-based dose-response curves were recently applied to the phytodosimetry of gamma radiation in a dicot plant, Arabidopsis thaliana, as an alternative biological assessment of genotoxicity using DNA damage response (DDR) genes. In the present study, we characterized gamma ray-responsive marker genes for transcriptome-based phytodosimetry in a monocot plant, rice (Oryza sativa L.), and compared different phytodosimetry models between rice and Arabidopsis using gamma-H2AX, comet, and quantitative transcriptomic assays. The transcriptome-based dose-response curves of four marker genes (OsGRG, OsMutS, OsRAD51, and OsRPA1) were reliably fitted to quadratic or exponential decay equations (r2 > 0.99). However, the single or integrated dose-response curves of these genes were distinctive from the conventional models obtained by the gamma-H2AX or comet assays. In comparison, rice displayed a higher dose-dependency in the comet signal and OsRAD51 transcription, while the gamma-H2AX induction was more dose-dependent in Arabidopsis. The dose-dependent transcriptions of the selected gamma-ray-inducible marker genes, including OsGRG, OsMutS, OsRAD51, and OsRPA1 in rice and AtGRG, AtPARP1, AtRAD51, and AtRPA1E in Arabidopsis, were maintained similarly at different vegetative stages. These results suggested that the transcriptome-based phytodosimetry model should be further corrected with conventional genotoxicity- or DDR-based models despite the high reliability or dose-dependency of the model. In addition, the relative weighting of each gene in the integrated transcriptome-based dose-response model using multiple genes needs to be considered based on the trend and amplitude of the transcriptional change.

Suppressive effect of tamarixetin, isolated from Inula japonica, on degranulation and eicosanoid production in bone marrow-derived mast cells.

BACKGROUND: The aim of this study was to evaluate the inhibitory effect of tamarixetin on the production of inflammatory mediators in IgE/antigen-induced mouse bone marrow-derived mast cells (BMMCs). MATERIALS AND METHODS: The effects of tamarixetin on mast cell activation were investigated with regard to degranulation, eicosanoid generation, Ca2+ influx, and immunoblotting of various signaling molecules. RESULTS: Tamarixetin effectively decreased degranulation and the eicosanoid generation such as leukotriene C4 and prostaglandin D2 in BMMCs. To elucidate the mechanism involved, we investigated the effect of tamarixetin on the phosphorylation of signal molecules. Tamarixetin inhibited the phosphorylation of Akt and its downstream signal molecules including IKK and nuclear factor κB. In addition, tamarixetin downregulated the phosphorylation of cytosolic phospholipase A2 (cPLA2) and p38 mitogen-activated protein kinase. CONCLUSIONS: Taken together, this study suggests that tamarixetin inhibits degranulation and eicosanoid generation through the PLCγ1 as well as Akt pathways in BMMCs, which would be potential for the prevention of allergic inflammatory diseases.

Effect of cilostazol on arteriovenous fistula in hemodialysis patients.

BACKGROUND: The maturation and patency of permanent vascular access are critical in patients requiring hemodialysis. Although numerus trials have been attempted to achieve permanently patent vascular access, little have been noticeable. Cilostazol, a phosphodiesterase-3 inhibitor, has been shown to be effective in peripheral arterial disease including vascular injury-induced intimal hyperplasia. We therefore aimed to determine the effect of cilostazol on the patency and maturation of permanent vascular access. METHODS: This single-center, retrospective study included 194 patients who underwent arteriovenous fistula surgery to compare vascular complications between the cilostazol (n=107) and control (n=87) groups. RESULTS: The rate of vascular complications was lower in the cilostazol group than in the control group (36.4% vs. 51.7%; p=0.033), including maturation failure (2.8% vs. 11.5%; p=0.016). The rate of reoperation due to vascular injury after hemodialysis initiation following fistula maturation was also significantly lower in the cilostazol group than in the control group (7.5% vs. 28.7%; p<0.001). However, there were no significant differences in the requirement for percutaneous transluminal angioplasty (PTA), rate of PTA, and the interval from arteriovenous fistula surgery to PTA between the cilostazol and control groups. CONCLUSION: Cilostazol might be beneficial for the maturation of permanent vascular access in patients requiring hemodialysis.

Effect of cilostazol on arteriovenous fistula in hemodialysis patients.

BACKGROUND: The maturation and patency of permanent vascular access are critical in patients requiring hemodialysis. Although numerus trials have been attempted to achieve permanently patent vascular access, little have been noticeable. Cilostazol, a phosphodiesterase-3 inhibitor, has been shown to be effective in peripheral arterial disease including vascular injury-induced intimal hyperplasia. We therefore aimed to determine the effect of cilostazol on the patency and maturation of permanent vascular access. METHODS: This single-center, retrospective study included 194 patients who underwent arteriovenous fistula surgery to compare vascular complications between the cilostazol (n=107) and control (n=87) groups. RESULTS: The rate of vascular complications was lower in the cilostazol group than in the control group (36.4% vs. 51.7%; p=0.033), including maturation failure (2.8% vs. 11.5%; p=0.016). The rate of reoperation due to vascular injury after hemodialysis initiation following fistula maturation was also significantly lower in the cilostazol group than in the control group (7.5% vs. 28.7%; p<0.001). However, there were no significant differences in the requirement for percutaneous transluminal angioplasty (PTA), rate of PTA, and the interval from arteriovenous fistula surgery to PTA between the cilostazol and control groups. CONCLUSION: Cilostazol might be beneficial for the maturation of permanent vascular access in patients requiring hemodialysis.

Flightless-1 inhibits ER stress-induced apoptosis in colorectal cancer cells by regulating Ca2+ homeostasis.

The endoplasmic reticulum (ER) stress response is an adaptive mechanism that is activated upon disruption of ER homeostasis and protects the cells against certain harmful environmental stimuli. However, critical and prolonged cell stress triggers cell death. In this study, we demonstrate that Flightless-1 (FliI) regulates ER stress-induced apoptosis in colon cancer cells by modulating Ca2+ homeostasis. FliI was highly expressed in both colon cell lines and colorectal cancer mouse models. In a mouse xenograft model using CT26 mouse colorectal cancer cells, tumor formation was slowed due to elevated levels of apoptosis in FliI-knockdown (FliI-KD) cells. FliI-KD cells treated with ER stress inducers, thapsigargin (TG), and tunicamycin exhibited activation of the unfolded protein response (UPR) and induction of UPR-related gene expression, which eventually triggered apoptosis. FliI-KD increased the intracellular Ca2+ concentration, and this upregulation was caused by accelerated ER-to-cytosolic efflux of Ca2+. The increase in intracellular Ca2+ concentration was significantly blocked by dantrolene and tetracaine, inhibitors of ryanodine receptors (RyRs). Dantrolene inhibited TG-induced ER stress and decreased the rate of apoptosis in FliI-KD CT26 cells. Finally, we found that knockdown of FliI decreased the levels of sorcin and ER Ca2+ and that TG-induced ER stress was recovered by overexpression of sorcin in FliI-KD cells. Taken together, these results suggest that FliI regulates sorcin expression, which modulates Ca2+ homeostasis in the ER through RyRs. Our findings reveal a novel mechanism by which FliI influences Ca2+ homeostasis and cell survival during ER stress.

PPM1A Controls Diabetic Gene Programming through Directly Dephosphorylating PPARγ at Ser273.

Peroxisome proliferator-activated receptor γ (PPARγ) is a master regulator of adipose tissue biology. In obesity, phosphorylation of PPARγ at Ser273 (pSer273) by cyclin-dependent kinase 5 (CDK5)/extracellular signal-regulated kinase (ERK) orchestrates diabetic gene reprogramming via dysregulation of specific gene expression. Although many recent studies have focused on the development of non-classical agonist drugs that inhibit the phosphorylation of PPARγ at Ser273, the molecular mechanism of PPARγ dephosphorylation at Ser273 is not well characterized. Here, we report that protein phosphatase Mg2+/Mn2+-dependent 1A (PPM1A) is a novel PPARγ phosphatase that directly dephosphorylates Ser273 and restores diabetic gene expression which is dysregulated by pSer273. The expression of PPM1A significantly decreases in two models of insulin resistance: diet-induced obese (DIO) mice and db/db mice, in which it negatively correlates with pSer273. Transcriptomic analysis using microarray and genotype-tissue expression (GTEx) data in humans shows positive correlations between PPM1A and most of the genes that are dysregulated by pSer273. These findings suggest that PPM1A dephosphorylates PPARγ at Ser273 and represents a potential target for the treatment of obesity-linked metabolic disorders.

Nepetin, a natural compound from Inulae flos, suppresses degranulation and eicosanoid generation through PLCγ1 and Akt signaling pathways in mast cells.

Nepetin derived from the flowers of Inula japonica, Inulae flos, has been reported to exert several biological activities, including anti-inflammatory responses. In this study, we evaluated the anti-allergic property of nepetin with its molecular mechanisms in bone marrow-derived mast cells (BMMC) and mice. In this in vitro study, we investigated the inhibitory effects of nepetin on degranulation and generation of leukotriene C4 (LTC4) and prostaglandin D2 (PGD2) in IgE/antigen (Ag)-stimulated BMMC. The effect of nepetin on passive cutaneous anaphylaxis (PCA) reaction was also studied in mice. Nepetin reduced degranulation and LTC4 generation in BMMC. The IgE/Ag-mediated signaling pathway demonstrated that nepetin suppressed intracellular Ca2+ level and activation of PLCγ1 and cPLA2. However, MAPKs were not affected by nepetin in BMMC. In addition, nepetin treatment reduced PGD2 production and suppressed cyclooxygenase-2 protein expression via the inhibition of the Akt and nuclear factor-κB signaling pathways. With respect to the local allergic response in vivo, oral administration of nepetin suppressed mast cell-dependent PCA reaction in a dose-dependent manner. The results of this study suggest that nepetin might have an anti-allergic potential related to mast cell-mediated inflammatory diseases.

Probability of causation for occupational cancer after exposure to ionizing radiation.

BACKGROUND: Probability of causation (PC) is a reasonable way to estimate causal relationships in radiation-related cancer. This study reviewed the international trend, usage, and critiques of the PC method. Because it has been used in Korea, it is important to check the present status and estimation of PC in radiation-related cancers in Korea. METHODS: Research articles and official reports regarding PC of radiation-related cancer and published from the 1980s onwards were reviewed, including studies used for the revision of the Korean PC program. PC has been calculated for compensation-related cases in Korea since 2005. RESULTS: The United States National Institutes of Health first estimated the PC in 1985. Among the 106 occupational diseases listed in the International Labor Organization Recommendation 194 (International Labor Office (ILO), ILO List of Occupational Diseases, 2010), PC is available only for occupational cancer after ionizing radiation exposure. The United States and United Kingdom use PC as specific criteria for decisions on the compensability of workers' radiation-related health effects. In Korea, PC was developed firstly as Korean Radiation Risk and Assigned Share (KORRAS) in 1999. In 2015, the Occupational Safety and Health Research Institute and Radiation Health Research Institute jointly developed a more revised PC program, Occupational Safety and Health-PC (OSH-PC). Between 2005 and 2015, PC was applied in 16 claims of workers' compensation for radiation-related cancers. In most of the cases, compensation was given when the PC was more than 50%. However, in one case, lower than 50% PC was accepted considering the possibility of underestimation of the cumulative exposure dose. CONCLUSIONS: PC is one of the most advanced tools for estimating the causation of occupational cancer. PC has been adjusted for baseline cancer incidence in Korean workers, and for uncertainties using a statistical method. Because the fundamental reason for under- or over-estimation is probably inaccurate dose reconstruction, a proper guideline is necessary.

A Vascular Endothelial Growth Factor-Dependent Sprouting Angiogenesis Assay Based on an In Vitro Human Blood Vessel Model for the Study of Anti-Angiogenic Drugs.

Angiogenesis is the formation of new capillaries from pre-existing blood vessels and participates in proper vasculature development. In pathological conditions such as cancer, abnormal angiogenesis takes place. Angiogenesis is primarily carried out by endothelial cells, the innermost layer of blood vessels. The vascular endothelial growth factor-A (VEGF-A) and its receptor-2 (VEGFR-2) trigger most of the mechanisms activating and regulating angiogenesis, and have been the targets for the development of drugs. However, most experimental assays assessing angiogenesis rely on animal models. We report an in vitro model using a microvessel-on-a-chip. It mimics an effective endothelial sprouting angiogenesis event triggered from an initial microvessel using a single angiogenic factor, VEGF-A. The angiogenic sprouting in this model is depends on the Notch signaling, as observed in vivo. This model enables the study of anti-angiogenic drugs which target a specific factor/receptor pathway, as demonstrated by the use of the clinically approved sorafenib and sunitinib for targeting the VEGF-A/VEGFR-2 pathway. Furthermore, this model allows testing simultaneously angiogenesis and permeability. It demonstrates that sorafenib impairs the endothelial barrier function, while sunitinib does not. Such in vitro human model provides a significant complimentary approach to animal models for the development of effective therapies.

A 3D in vitro pericyte-supported microvessel model: visualisation and quantitative characterisation of multistep angiogenesis.

Angiogenesis, which refers to the formation of new blood vessels from already existing vessels, is a promising therapeutic target and a complex multistep process involving many different factors. Pericytes (PCs) are attracting attention as they are considered to make significant contributions to the maturation and stabilisation of newly formed vessels, although not much is known about the precise mechanisms involved. Since there is no single specific marker for pericytes, in vivo models may complicate PC identification and the study of PCs in angiogenesis would benefit from in vitro models recapitulating the interactions between PCs and endothelial cells (ECs) in a three-dimensional (3D) configuration. In this study, a 3D in vitro co-culture microvessel model incorporating ECs and PCs was constructed by bottom-up tissue engineering. Angiogenesis was induced in the manner of sprout formation by the addition of a vascular endothelial cell growth factor. It was found that the incorporation of PCs prevented expansion of the parent vessel diameter and enhanced sprout formation and elongation. Physical interactions between ECs and PCs were visualised by immunostaining and it disclosed that PCs covered the EC monolayer from its basal side in the parent vessel as well as angiogenic sprouts. Furthermore, the microvessels were visualized in 3D by using a non-invasive optical coherence tomography (OCT) imaging system and sprout features were quantitatively assessed. It revealed that the sprouts in EC-PC co-culture vessels were longer and tighter than those in EC mono-culture vessels. The combination of the microvessel model and the OCT system analysis can be useful for the visualisation and demonstration of the multistep process of angiogenesis, which incorporates PCs.

Anti-inflammatory Effects of KOTMIN13: A Mixed Herbal Medicine in LPS-stimulated RAW 264.7 Cells and Mouse Edema Models.

BACKGROUND: A Korean herbal medicine, KOTMIN13, composed of Inula japonica Thunberg, Trichosanthes kirilowii Maximowicz var. japonica kitamura, Peucedanum praeruptorum Dunn, and Allium macrostemon Bge, has been used for anti-allergic and anti-asthmatic treatment in oriental clinics, but its activity has not been investigated. MATERIALS AND METHODS: To evaluate the anti-inflammatory activity of KOTMIN13 for in vitro study, LPS-stimulated RAW 264.7 cells were used to induce the production and expression of inflammatory mediators and its mechanisms. 12-O-Tetradecanoylphorobol-13 aceate (TPA)-induced ear edema and carrageenan-induced paw edema models were also used to evaluate the effect of KOTMIN13 on acute inflammation in vivo. RESULTS: KOTMIN13 reduced the release of inflammatory mediators [nitric oxide, prostaglandin E2, interleukin (IL)-1ß, and IL-6] and the protein expression of inducible nitric oxide synthase and cyclooxygenase-2 in LPS-stimulated RAW 264.7 cells. Mechanism studies showed the attenuation of LPS-induced NF-κB activation by KOTMIN13 via IκBα degradation abrogation and a subsequent decrease in nuclear p65 levels. Activation of mitogen-activated protein kinases (ERK, JNK, and p38) was also suppressed. Furthermore, KOTMIN13 ameliorated the development of TPA-induced ear edema and carrageenan-induced paw edema in acute inflammatory edema mouse models. CONCLUSION: Our study demonstrates that KOTMIN13 inhibits inflammatory mediators through the inhibitions of NF-κB and MAPK activities in LPS-induced RAW 264.7 cells, as well as acute inflammation in edema models, indicating that KOTMIN13 is an effective suppressor for anti-inflammatory activities. SUMMARY: KOTMIN13 decrease the production of No, PGE2, and proinflammatory cytokine (TNF-∝, IL-1ß,IL-6).KOTMIN13 Suppressed the degradation of NF-kß and IKßα and the phosorylation of MAP Kinases.Topical application of KOTMIN13 reduced mouse ear edema.Oral administration of KOTMIN13 decreased carrageenan-induced paw edema. Abbreviations used: NO: nitric oxide; PGE2: prostaglandin E2; iNOS: inducible NO synthase; COX-2: cyclooxygenase-2; TNF-α: tumor necrosis factor-α; IL: interleukin; NF-κB: nuclear factor kappaB; MAPK: mitogen-activated protein kinases; ERK: extracellular signal regulated kinase; JNK: c-jun N terminal kinase; TPA: 12-O-tetradecanoylphorbol-13-acetate.

Methionine sulfoxide reductase B1 deficiency does not increase high-fat diet-induced insulin resistance in mice.

Methionine-S-sulfoxide reductase (MsrA) protects against high-fat diet-induced insulin resistance due to its antioxidant effects. To determine whether its counterpart, methionine-R-sulfoxide reductase (MsrB) has similar effects, we compared MsrB1 knockout and wild-type mice using a hyperinsulinemic-euglycemic clamp technique. High-fat feeding for eight weeks increased body weights, fat masses, and plasma levels of glucose, insulin, and triglycerides to similar extents in wild-type and MsrB1 knockout mice. Intraperitoneal glucose tolerance test showed no difference in blood glucose levels between the two genotypes after eight weeks on the high-fat diet. The hyperglycemic-euglycemic clamp study showed that glucose infusion rates and whole body glucose uptakes were decreased to similar extents by the high-fat diet in both wild-type and MsrB1 knockout mice. Hepatic glucose production and glucose uptake of skeletal muscle were unaffected by MsrB1 deficiency. The high-fat diet-induced oxidative stress in skeletal muscle and liver was not aggravated in MsrB1-deficient mice. Interestingly, whereas MsrB1 deficiency reduced JNK protein levels to a great extent in skeletal muscle and liver, it markedly elevated phosphorylation of JNK, suggesting the involvement of MsrB1 in JNK protein activation. However, this JNK phosphorylation based on a p-JNK/JNK level did not positively correlate with insulin resistance in MsrB1-deficient mice. Taken together, our results show that, in contrast to MsrA deficiency, MsrB1 deficiency does not increase high-fat diet-induced insulin resistance in mice.

KOTMIN13, a Korean herbal medicine alleviates allergic inflammation in vivo and in vitro.

BACKGROUND: The ethanol extract of KOTMIN13, composed of Inula japonica Flowers, Trichosanthes kirilowii Semen, Peucedanum praeruptorum Radix, and Allium macrostemon Bulbs, was investigated for its anti-asthmatic and anti-allergic activities. METHODS: The anti-asthmatic effects of KOTMIN13 were evaluated on ovalbumin (OVA)-induced murine asthma model. Anti-allergic properties of KOTMIN13 in bone-marrow derived mast cells (BMMC) and passive cutaneous anaphylaxis (PCA) in vivo were also examined. RESULTS: In asthma model, KOTMIN13 effectively suppressed airway hyperresponsiveness induced by aerosolized methacholine when compared to the levels of OVA-induced mice. KOTMIN13 treatment reduced the total leukocytes, eosinophil percentage, and Th2 cytokines in the bronchoalveolar lavage fluids in OVA-induced mice. The increased levels of eotaxin and Th2 cytokines in the lung as well as serum IgE were decreased by KOTMIN13. The histological analysis shows that the increased inflammatory cell infiltration and mucus secretion were also reduced. In addition, the degranulation and leukotriene C4 production were inhibited in BMMC with IC50 values of 3.9 µg/ml and 1.7 µg/ml, respectively. Furthermore, KOTMIN13 treatment attenuated mast-mediated PCA reaction. CONCLUSIONS: These results demonstrate that KOTMIN13 has anti-asthmatic and anti-allergic effects in vivo and in vitro models.

Britanin attenuates ovalbumin-induced airway inflammation in a murine asthma model.

We previously demonstrated the alleviation of ovalbumin (OVA)-induced airway inflammation by Inulae flos. In the present study, the effects of britanin, a sesquiterpene compound isolated from Inulae flos, were evaluated in an in vivo animal model for anti-asthma activity through observation of airway hyperresponsiveness (AHR), eosinophil recruitment, Th2 cytokine and IgE levels, and lung histopathology. Britanin administration effectively reduced AHR induced by aerosolized methacholine, airway eosinophilia, Th2 cytokines in bronchoalveolar lavage fluids and the supernatant of cultured splenocytes compared with OVA-induced mice. Histological studies showed that increased inflammatory cell infiltration and mucus secretion were reduced by britanin administration. Thus, britanin may have therapeutic potential for treating allergic asthma.