The application of augmented reality for improving clinical skills: a scoping review.

Augmented reality technology had developed rapidly in recent years and had been applied in many fields, including medical education. Augmented reality had potential to improve students' knowledge and skills in medical education. This scoping review primarily aims to further elaborate the current studies on the implementation of augmented reality in advancing clinical skills. This study was conducted by utilizing electronic databases such as PubMed, Embase, and Web of Science in June 2022 for articles focusing on the use of augmented reality for improving clinical skills. The Rayyan website was used to screen the articles that met the inclusion criteria, which was the application of augmented reality as a learning method in medical education. Total of 37 articles met the inclusion criteria. These publications suggested that using augmented reality could improve clinical skills. The most researched topics explored were laparoscopic surgery skills and ophthalmology were the most studied topic. The research methods applied in the articles fall into two main categories: randomized control trial (RCT) (29.3%) and non-RCT (70.3%). Augmented reality has the potential to be integrated in medical education, particularly to boost clinical studies. Due to limited databases, however, any further studies on the implementation of augmented reality as a method to enhance skills in medical education need to be conducted.

Hypolipidemic effect of brown seaweed (Sargassum crassifolium) extract in vivo (Study of histopathology, mRNA expression, and immunohistochemistry (IHC) with VCAM-1, ICAM-1, and MCP-1 parameters).

The purpose of this study was to obtain natural drugs from brown seaweed (Sargassum crassifolium) as antiatherosclerosis candidates through the study of hypolipidemic mechanisms of action. Modeling of dyslipidemia rats was carried out by feeding high-fat (HFF) and doses of crude fucoidan 100. 200. 400mg / KgBB. in both treatments measured blood lipid profile levels taken from the orbital sinuses. HE's histopathology. mRNA expression. immunohistochemistry (IHC) with parameters VCAM-1. ICAM-1. and MCP-1 were performed on adipose tissue. as well as liver. Total cholesterol values 51.07-225.2. triglycerides 30.43-115.73. HDL 13.1-24.86 mg/dl and LDL 20.22-189.68 mg/dl. In the treatment of crude fucoidan obtained the result of p value < α (0.05. Histopathological features of adipose tissue after administration of HFF for 60 days resulted in an increase in adipose cell size. and the liver experienced structural damage and inflammation. but after 21 days of treatment the morphological picture of adipose tissue was similar to normal morphology and the liver also decreased in severity and inflammation. The results of histochemical staining after treatment showed a positive staining part on MCP-1. The result of p value < α (0.05) of mRNA expression for administration of 3 treatment doses. A dyslipidemic mouse model with HFF administration for 60 days succeeded in becoming a dyslipidemic rat. and crude fucoidan had hypolipidemic activity. Doses of 100. 200. and 400 mg/KgBB crude fucoidan showed improvement in adipose and liver morphological features of severity and inflammation of dyslipidemic rats and decreased mRNA expression.

Matrix Metalloproteinase-9 Testing of Golden Rice Cookies With Piper Crocatum Active Extract for Preventing Foot Ulcers in Patients With Diabetes: Protocol for a Randomized Controlled Trial.

BACKGROUND: Diabetic foot ulcers (DFUs) present a formidable challenge to both patients and health care systems. DFUs significantly reduce the quality of life for patients, prolong hospital stays, and are the cause of approximately 70,000 lower limb amputations across the globe annually. Prevention of DFUs primarily involves the optimization of blood sugar levels and the effective management of complications, particularly peripheral neuropathy. Golden Rice has been proven to lower blood sugar levels due to its beta-carotene content, and Piper crocatum (P. crocatum) has been found to be effective in reducing the risk factors of DFUs through biomolecular regulation because of its polyphenol content. OBJECTIVE: The principal objective of this study is to identify the efficacy of P. crocatum-enriched cookies, with Golden Rice as their primary ingredient, in preventing DFUs. The evaluation will center on their impact on the expression of matrix metalloproteinase-9 (MMP-9), a pivotal factor in the development of DFUs. METHODS: This study is an experimental clinical research that follows the randomized controlled trial method and uses a single-blind design. The participants in the study are outpatients from primary health centers in Makassar, Indonesia, who have been diagnosed with diabetes mellitus. The sample for the study will be randomly selected and subsequently categorized into 2 groups: the intervention group and the control group. The intervention group consumes P. crocatum-enriched Golden Rice cookies, while the control group receives cookies without these additives. The participants from both groups will consume their respective cookies (packaged identically) twice a day for 14 days. The cookies will be prepared according to a modified recipe with an emphasis on low glucose content, resulting in 51 calories per cookie, comprising 1% carbohydrates, 6% fat, 4% cholesterol, and 4% fiber, excluding gluten, sugar, and salt. They will be baked at 158°C for 20 minutes. The process involves the addition of 20% Golden Rice and 10% P. crocatum ethanol extract, both prepared via maceration with 96% ethanol. The dependent variable in this study is the expression of gelatinases matrix metalloproteinase, to be assessed at 2 distinct time points-preintervention (pretest) and postintervention (posttest)-with the evaluation conducted through the western blotting method. RESULTS: The recruitment and testing phase started in January 2024. The study is scheduled to be completed by the end of March 2024. Data analysis will commence in April 2024, and the publication of the results is anticipated in the same year (2024). The study will report on the changes in primary data, encompassing gelatinases matrix metalloproteinase, as well as secondary data, including the ankle-brachial index, neuropathy score, and random blood glucose level. CONCLUSIONS: The findings of this trial are expected to significantly impact the selection of strategies by health care practitioners to enhance diabetes self-management, particularly in the domain of therapeutic snacking, for patients diagnosed with diabetes mellitus. TRIAL REGISTRATION: Thai Clinical Trials Registry TCTR20230502001; https://www.thaiclinicaltrials.org/show/TCTR20230502001. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): PRR1-10.2196/49940.

Chlorogenic Acid Inhibits Progressive Pulmonary Fibrosis in a Diabetic Rat Model.

Background: Chlorogenic acid (CGA) is known to have antifibrotic and hypoglycemic effects and may play a role in preventing diabetes-induced pulmonary fibrosis. This study aimed to determine the effect and optimum dose of CGA on diabetes-induced pulmonary fibrosis. Methods: Thirty Wistar rats (two-month-old, 150-200 grams) were randomly divided into six groups, namely control, six weeks diabetes mellitus (DM1), eight weeks DM (DM2), and three DM2 groups (CGA1, CGA2, and CGA3) who received CGA doses of 12.5, 25, and 50 mg/Kg BW, respectively. After six weeks, CGA was administered intraperitoneally for 14 consecutive days. Lung tissues were taken for TGF-ß1, CTGF, SMAD7, Collagen-1, and α-SMA mRNA expression analysis and paraffin embedding. Data were analyzed using one-way ANOVA and the Kruskal-Wallis test. P<0.05 was considered statistically significant. Results: TGF-ß1 expression in the CGA1 group (1.01±0.10) was lower than the DM1 (1.33±0.25, P=0.05) and DM2 (1.33±0.20, P=0.021) groups. α-SMA expression in the CGA1 group (median 0.60, IQR: 0.34-0.64) was lower than the DM1 (median 0.44, IQR: 0.42-0.80) and DM2 (median 0.76, IQR: 0.66-1.10) groups. Collagen-1 expression in the CGA1 group (0.75±0.13) was lower than the DM1 (P=0.24) and DM2 (P=0.26) groups, but not statistically significant. CTGF expression in CGA groups was lower than the DM groups (P=0.088), but not statistically significant. There was an increase in SMAD7 expression in CGA groups (P=0.286). Histological analysis showed fibrosis improvement in the CGA1 group compared to the DM groups. Conclusion: CGA (12.5 mg/Kg BW) inhibited the expression of profibrotic factors and increased antifibrotic factors in DM-induced rats.

The effect of preconditioning hypoxia in schwann-like-cells-derived adipose mesenchymal stem cells and rat sciatic nerve-derived stem cells: experimental research.

The preconditioning hypoxia for stem cells is a strategy to achieve effective conditions for cell therapy, indicate increased expression of regenerative genes in stem cell therapy, and enhance the secretion of bioactive factors and therapeutic potential of their cultured secretome. Objectives: This study aims to explore the response of Schwann-like cells derived from adipose-derived mesenchymal stem cells (SLCs) and Schwann cells rat sciatic nerve-derived stem cells (SCs) with their secretomes under normoxic and hypoxic conditions in vitro. Material and methods: SLCs and SCs were isolated from the adipose tissue and the sciatic nerve of the adult white male rat strain Wistar. Cells were incubated in 21% O2 (normoxic group) and 1%, 3%, and 5% O2 (hypoxic group) conditions. Concentration values of transforming growth factor-ß (TGF-ß), basic Fibroblast Growth factor (bFGF), brain-derived neurotrophic factor, glial-derived neurotrophic factor, vascular endothelial growth factor, and nerve growth factor were detected and calculated utilizing an enzyme-linked immunosorbent assay, and the growth curve was described. Results: SLCs and SCs indicated positive expression for mesenchymal markers and negative expression for hematopoietic markers. Normoxic conditions SLCs and SCs showed elongated and flattened morphology. Under hypoxic conditions, SLCs and SCs showed a classic fibroblast-like morphology. Hypoxia 1% gave the highest concentration in TGF-ß and bFGF from the SLCs group and TGF-ß, bFGF, brain-derived neurotrophic factor, and vascular endothelial growth factor from the SCs group. No significant differences in concentration of growth factors between the SLCs group compared to SCs group in all oxygen groups. Conclusions: Preconditioning hypoxia has an effect on the composing of SLCs, SCs, and their secretomes in vitro; no significant differences in concentration of growth factors between the SLCs group compared with the SCs group in all oxygen groups.

The optimum oxygen level in hypoxic culture conditions of ligament derived stem cells: experimental research.

The hypoxic condition is a physiological norm for various stem cells. The natural microenvironment contains lower oxygen pressures. Recent studies reported significant increases in the cultured cells' proliferation in the presence of a low oxygen pressure. Objective: This study aimed to investigate the optimum oxygen level for rabbit cruciate ligament fibroblast cells culture and Ligament Derived Conditioned Medium/LD-CM (Secretome) preparation in vitro. Materials and methods: Fibroblasts were isolated from the cruciate ligament of the rabbit's knee. Cultured of rabbit cruciate ligament Fibroblast Cells (fifth passage) were assigned to the slight (5% O2), middle (3% O2), and severe hypoxia (1% O2) groups and the normoxia (21% O2) group. Measurement of growth factors: TGF-ß1, PDGF, FGF, and VEGF in LD-CM (Secretome) used an enzyme-linked immunosorbent assay. Results: The highest number of cultured cells were in the 5% O2 group compared to the normoxia, 1 and 3% groups. The hypoxia 5% group also had increased productions of PDGF, FGF, and VEGF proteins in LD-CM (secretome) compared to the 1, 3%, and normoxia groups. TGF-ß1 production was slightly higher in the 3 group than the 5% group. Conclusion: The hypoxic precondition of 5% oxygen was the optimum condition for ligament culture and ligament derived conditioned medium (secretome) preparation in vitro.

Calcitriol attenuates vascular remodeling in association with alteration of ppET-1/ETBR/eNOS and ETAR expression in acute and chronic phases of kidney ischemia-reperfusion injury in mice.

Kidney ischemia-reperfusion injury (IRI) causes acute kidney injury with increasing risk of maladaptive repair through endothelin-1 (ET-1)/endothelin type A receptor (ETAR) signaling. Calcitriol shows renoprotection in kidney fibrosis, however, its effects on vasoactive substances expression and vascular remodeling following kidney IRI remain unclear. This research aimed to investigate Calcitriol's effects on preproendothelin-1 (ppET-1), ETAR, endothelial nitric oxide synthase (eNOS) mRNA expression and vascular remodeling in acute and chronic phases of kidney IRI in mice. Twenty-five male Swiss mice were randomly divided into five groups (n = 5): SO (sham-operated), IR3 (3 day kidney IRI), IR12 (12 day kidney IRI), IRD3 (3 day kidney IRI + Calcitriol 0.5 µg/kg body weight (BW)/day), and IRD12 (12 day kidney IRI + Calcitriol 0.5 µg/kg BW/day). Ischemia-reperfusion injury groups underwent bilateral renal pedicles clamping for 30 min, then reperfusion. Kidneys were harvested for Sirius Red staining to observe interstitial fibrosis and vascular remodeling, polymerase chain reaction to quantify ppET-1, endothelin type B receptor (ETBR), eNOS mRNA expression, and Western blotting to quantify ETAR protein expression. Calcitriol treatment in both phases of kidney IRI showed lower serum creatinine and ETAR protein expression, while higher eNOS and ETBR mRNA expression than IRI-only groups. Furthermore, ppET-1 mRNA expression was higher in IRD3 than IR3, but lower in IRD12 than IR12. Calcitriol also prevented vascular remodeling as indicated by lower wall thickness and higher lumen/wall area ratio than IRI-only groups.

Curcumin Prevents Epithelial-to Mesenchymal Transition-Mediated Ovarian Cancer Progression through NRF2/ETBR/ET-1 Axis and Preserves Mitochondria Biogenesis in Kidney after Cisplatin Administration.

Purpose: Ovarian carcinoma is one of the gynaecological malignancies that have the highest mortality rates due to its progressivity. Endothelin signalling plays a leading role in the progression of ovarian cancer through Epithelial-to-Mesenchymal Transition (EMT). Cisplatin commonly used as potent chemotherapy; however, its application hindered by its nephrotoxic effect. Curcumin, a turmeric-derived compound, has an anticancer property, as well as a renal protective effect. Moreover, curcumin augments the affinity of the antioxidant enzyme, while inhibits endothelin-1 (ET-1) signalling. The effects of curcumin on ovarian cancer progression and cisplatin-induced kidney injury remain unknown. Methods: Curcumin was used as a supplementary therapy together with cisplatin in Human Ovarian Cancer Cell line (SKOV3) and also in rodent-induced ovarian cancer. The kidney phenotype in the ovarian cancer rat model after cisplatin ± curcumin administration will also be analyzed Results: Co-treatment of cisplatin with curcumin enhanced the expression of a gene involved in apoptosis in association with NRF2 enhancement, thus activated ETBR-mediated ET-1 clearance in SKOV3 cell and ovarian cancer model in rat. Moreover, curcumin treatment improved mitochondria biogenesis markers such as PGC-1α and TFAM and prevented the elevated of ET-1-mediated renal fibrosis and apoptosis in kidney isolated from cisplatin-treated ovarian cancer rat. Conclusion: Curcumin could be potentially added as an anticancer adjuvant with protective effects in the kidney; thus, improves the efficacy and safety of cisplatin treatment in the clinical setting.

Accelerated Senescence and Apoptosis in the Rat Liver during the Progression of Diabetic Complications.

Background: Chronic hyperglycaemia of diabetes causes long-term damage and impaired function of multiple organs. However, the pathological changes in the liver following long-term diabetes remain unclear. This study aimed to determine the pathological complications of long-term diabetes in the rat liver. Methods: Intraperitoneal injection of streptozotocin (STZ) was used to induce diabetes in rats at a single dose (60 mg/kg body weight [BW]). Rats were euthanised at 1 month (DM1 group), 2 months (DM2 group) and 4 months (DM4 group) following diabetes induction with six rats in each group. Immunohistochemistry was performed against SOD1, CD68, p53 and p16 antibodies. Messenger RNA (mRNA) expressions of SOD1, SOD2, GPx, CD68, p53, p21 and caspase-3 genes were measured by reverse transcription-polymerase chain reaction. Results: Hepatic p53 mRNA expression was significantly higher in DM1, DM2 and DM4 groups compared to the control group. The p21 and caspase-3 mRNA expressions were significantly upregulated in the DM2 and DM4 groups. The p16-positive cells were obviously increased, particularly in the DM4 group. Bivariate correlation analysis showed mRNA expressions of p21 and caspase-3 genes were positively correlated with the p53 gene. Conclusion: Diabetic rats exhibited increased apoptosis and senescence in the liver following a longer period of hyperglycaemia.

Ethanolic Extract of Centella asiatica Treatment in the Early Stage of Hyperglycemia Condition Inhibits Glomerular Injury and Vascular Remodeling in Diabetic Rat Model.

BACKGROUND: Diabetes mellitus (DM) is marked by oxidative stress, inflammation, and vascular dysfunction that caused diabetic nephropathy that resulted in end-stage renal disease (ESRD). Vascular dysfunction is characterized by an imbalance in vasoconstrictor and vasodilator agents which underlies the mechanism of vascular injury in DM. Additionally, diminished podocytes correlate with the severity of kidney injury. Podocyturia often precedes proteinuria in several kidney diseases, including diabetic kidney disease. Centella asiatica (CeA) is known as an anti-inflammatory and antioxidant and has neuroprotective effects. This research aimed to investigate the potential effect of CeA to inhibit glomerular injury and vascular remodeling in DM. METHODS: The DM rat model was induced through intraperitoneal injection of streptozotocin 60 mg/kg body weight (BW), and then rats were divided into 1-month DM (DM1, n = 5), 2-month DM (DM2, n = 5), early DM concurrent with CeA treatment for 2 months (DMC2, n = 5), and 1-month DM treated with CeA for 1-month (DM1C1, n = 5). The CeA (400 mg/kg BW) was given daily via oral gavage. The control group (Control, n = 5) was maintained for 2 months. Finally, rats were euthanized and kidneys were harvested to assess vascular remodeling using Sirius Red staining and the mRNA expression of superoxide dismutase, podocytes marker, ACE2, eNOS, and ppET-1 using RT-PCR. RESULTS: The DM groups demonstrated significant elevation of glucose level, glomerulosclerosis, and proteinuria. A significant reduction of SOD1 and SOD3 promotes the downregulation of nephrin and upregulation of TRPC6 mRNA expressions in rat glomerular kidney. Besides, this condition enhanced ppET-1 and inhibited eNOS and ACE2 mRNA expressions that lead to the development of vascular remodeling marked by an increase of wall thickness, and lumen wall area ratio (LWAR). Treatment of CeA, especially the DMC2 group, attenuated glomerular injury and showed the reversal of induced conditions. CONCLUSIONS: Centella asiatica treatment at the early stage of diabetes mellitus ameliorates glomerulosclerosis and vascular injury via increasing antioxidant enzymes.

Centella asiatica Extract Attenuates Kidney Fibrosis Through Reducing Mesenchymal Transition and Inflammation in Ureteral Ligation Model in Mice.

Background: Kidney fibrosis is the common final pathway of chronic kidney disease (CKD), and is characterized by inflammation, mesenchymal transition with myofibroblast formation and epithelial to mesenchymal transition (EMT). Centella asiatia (CeA) is an herb that has a reno-protective effect. However, its mechanism of action in kidney fibrosis has not been elucidated. Aim: To elucidate the effect of CeA in amelioration of kidney fibrosis in a unilateral ureteral obstruction (UUO) model and focus on mesenchymal transition and inflammation. Methods: Unilateral ureteral obstruction was performed in male Swiss-background mice (age: 2-3 months, weight: 30-40 g, UUO group n = 6) to induce kidney fibrosis. Two doses of CeA extract with oral administration, 210 and 840 mg/kg body weight were added in UUO (U+C210 and U+C840 groups, each n = 6). The sham operation procedure was performed for the control group (SO, n = 6). The mice were euthanized at day-14 after operation. Tubular injury and interstitial fibrosis area fractions in kidney tissues of the mice were quantified based on periodic acid-Schiff (PAS) and Sirius Red (SR) staining. Immunostaining was performed for examination of fibroblast (PDGFR-ß), myofibroblast (α-SMA), Monocyte Chemoattractant Protein-1 (MCP-1) and macrophage (CD68), meanwhile double immunofluorescence was performed with PDGFR-ß and α-SMA. Reverse transcriptase-polymerase chain reaction (RT-PCR) was performed to examine mRNA expression of TGF-ß, Collagen-1, Snail, E-cadherin, vimentin, fibroblast-specific protein 1 (FSP-1), CD68, toll-like receptor 4 (TLR4), and MCP-1. Results: We observed a significantly higher interstitial fibrosis area fraction and tubular injury (p < 0.001) with fibroblast expansion and myofibroblast formation in the UUO group than in the SO group. These findings were associated with higher mRNA expression of TGF-ß, Collagen-1, Snail, vimentin, FSP-1, CD68, TLR4, and MCP-1 and lower mRNA expression of E-cadherin. The U+C840 group had a significantly lower tubular injury score and interstitial fibrosis area fraction, which associated with downregulation of mRNA expression of TGF-ß, Collagen-1, Snail, vimentin, FSP-1, CD68, TLR4, and MCP-1, with upregulation of mRNA expression of E-cadherin. Immunostaining observation revealed the U+C840 group demonstrated reduction of macrophage infiltration and myofibroblast expansion. Conclusion: CeA treatment with dose-dependently ameliorates mesenchymal transition and inflammation in kidney fibrosis in mice.

Effect of semaphorin 3C gene variants in multifactorial Hirschsprung disease.

OBJECTIVE: Cluster genes, specifically the class 3 semaphorins (SEMA3) including SEMA3C, have been associated with the development of Hirschsprung disease (HSCR) in Caucasian populations. We aimed to screen for rare and common variants in SEMA3C in Indonesian patients with HSCR. METHODS: In this prospective clinical study, we analyzed SEMA3C gene variants in 55 patients with HSCR through DNA sequencing and bioinformatics analyses. RESULTS: Two variants in SEMA3C were found: p.Val337Met (rs1527482) and p.Val579 = (rs2272351). The rare variant rs1527482 (A) was significantly overrepresented in our HSCR patients (9.1%) compared with South Asian controls in the 1000 Genomes (4.7%) and Exome Aggregation Consortium (ExAC; 3.5%) databases. Our analysis using bioinformatics tools predicted this variant to be evolutionarily conserved and damaging to SEMA3C protein function. Although the frequency of the other variant, rs2272351 (G), also differed significantly in Indonesian patients with HSCR (27.3%) from that in South Asian controls in 1000 Genomes (6.2%) and ExAC (4.6%), it is a synonymous variant and not likely to affect protein function. CONCLUSIONS: This is the first comprehensive report of SEMA3C screening in patients of Asian ancestry with HSCR and identifies rs1527482 as a possible disease risk allele in this population.

The impact of COL1A1 and COL6A1 expression on hypospadias and penile curvature severity.

BACKGROUND: Hypospadias, the most frequent congenital male external genitalia abnormality, is usually associated with curvature of the ventral penis, i.e. chordee. Abnormality of darto tissue has been suggested as the pathophysiology of chordees. Collagen is one of the most abundant fibrous proteins within the extracellular matrix. In this study, we determined the expression of collagen 1 (COL1A1) and COL6A1 in patients with hypospadias and associated them with the severity of penile curvature. METHODS: We included 60 children < 18 years old, consisting of 20 distal hypospadias, 20 proximal hypospadias patients, and 20 controls in our institution from 2017 - 2020. The expression of COL1A1 and COL6A1 in darto tissue was determined by reverse-transcriptase polymerase chain reaction (qPCR). The penile curvature severity was classified as mild (< 30 degrees), moderate (30-60 degrees), and severe (> 60 degrees). RESULTS: qPCR showed that COL1A1 and COL6A1 expression was significantly downregulated in the distal (0.88 (0.38-2.53) and 0.54 (0.16-4.35), respectively) and proximal 0.76 (0.33-2.57) and 0.57 (0.18-1.38), respectively) hypospadias groups compared to controls (1.85 (0.24-4.61) and 0.93 (0.17-4.06), respectively) with p-values of 0.024 and 0.018, respectively. Furthermore, there was a moderate correlation between COL1A1 and COL6A1 expression (r = 0.458, p < 0.0001). Interestingly, COL1A1 and COL6A1 were also significantly downregulated in the moderate and severe chordee groups compared to the mild chordee groups, with p-values of 0.003 and 0.037, respectively. CONCLUSIONS: Aberrant COL1A1 and COL6A1 expression might affect abnormalities in darto tissue and penile curvature severity in hypospadias patients.

Upregulation of p16, Bax and Bcl-2 mRNA Expression Associated with Epithelial Apoptosis and Myofibroblast Proliferation in Kidney Fibrosis Model in Mice.

BACKGROUND: Cellular senescence may play a role in the development of kidney fibrosis, but its specific association with apoptosis or proliferation have yet to be determined. OBJECTIVES: This study aims to determine the effects of unilateral ureteral obstruction (UUO) on proliferation, cellular senescence and apoptosis in kidney fibrosis. METHODS: A unilateral ureteral obstruction (UUO) procedure was performed to induce kidney fibrosis in 24 Swiss mice (3 months old, 30 g-40 g). Mice were sacrificed on day 3 (UUO3, n = 6), day 7 (UUO7, n = 6) and day 14 (UUO14, n = 6). Sham operation (SO) procedures were performed on the control group. The expression of Bcl-2, p16 and Bax mRNA was quantified with reverse transcription polymerase chain reaction (RT-PCR). Immunohistochemical (IHC) staining with anti-Bcl-2 and p53 antibodies was used to determine the localisation of proliferation and apoptosis. Data were analysed using one-way ANOVA followed by a post hoc least significant difference (LSD) test (P < 0.05). RESULTS: RT-PCR analysis showed higher mRNA expression of Bcl-2, p16 and Bax in the UUO groups compared with SO group (P < 0.05). Immunostaining showed that Bcl-2 and p53 expression in tubular epithelium in the UUO groups, except Bcl-2 expression was found in interstitial areas of UUO14 group. CONCLUSION: Senescence in UUO might be associated with epithelial apoptosis and myofibroblast proliferation.

Endothelin converting enzyme-1 (ECE-1) deletion in association with Endothelin-1 downregulation ameliorates kidney fibrosis in mice.

Kidney fibrosis is a common final pathway of chronic kidney diseases, which are characterized by renal architecture damage, inflammation, fibroblast expansion and myofibroblast formation. Endothelin converting enzyme-1 (ECE-1) contributes to activation of Endothelin-1 (ET-1), a potent vasoconstrictor and pro-fibrotic substance. This study elucidated the effect of ECE-1 knockout in kidney fibrosis model in mice in association of ET-1 downregulation. Kidney fibrosis was performed in ECE-1 knockout (ECE-1 KO) and vascular endothelial derived ET-1 KO (VEETKO) mice (2 months, 20-30 g, n = 30) and their wild type (WT) littermates using unilateral ureteral obstruction (UUO) procedure. Mice were euthanized on day-7 and day-14 after UUO. Histopathological analysis was conducted for fibrosis and tubular injury. Immunostainings were done to quantify macrophages (F4/80), fibroblasts (FSP-1) and myofibroblasts (α-SMA). Monocyte Chemoattractant Protein-1 (MCP-1), ECE-1 and preproET-1 (ppET-1) mRNA expression were quantified with qRT-PCR, while Transforming Growth Factor-ß1 (TGF-ß1) and α-SMA protein level were quantified with Western blot. ECE-1 KO mice demonstrated reduction of ECE-1 and ppET-1 mRNA expression, attenuation of kidney fibrosis, tubular injury, MCP-1 mRNA expression and macrophage number compared to WT. Double immunostaining revealed fibroblast to myofibroblast formation after UUO, while ECE-1 KO mice had significantly lower fibroblast number and myofibroblast formation compared to WT, which were associated with significantly lower TGF-ß1 and α-SMA protein levels in day-14 of UUO. VEETKO mice also demonstrated attenuation of ET-1 protein level, fibrosis and myofibroblast formation. In conclusion, ECE-1 knockout and ET-1 downregulation attenuated kidney fibrosis.

Achatina fulica mucous improves cell viability and increases collagen deposition in UVB-irradiated human fibroblast culture.

Introduction: Ultraviolet radiation induces skin photoaging by increasing matrix metalloproteinase-1 (MMP-1). MMP-1 degrades type I and III collagen that comprise the dermal connective tissue. Achatina fulica mucous (AFM) is a natural remedy that has protective effects on fibroblasts and collagen. Objective: To investigate the effects of AFM on cell viability and collagen deposition in UVB-irradiated human fibroblast culture. Methods: The mucous was extracted from 50 Achatina fulica snails that were stimulated by a 5-10 Volt electricity shock for 30-60 seconds and converted into powder by the freeze-drying process. The human dermal fibroblast culture was divided into six groups: group 1 were normal fibroblasts without UVB irradiation as normal control, groups 2-5 consisted of 100 mJ/cm2 UVB-irradiated fibroblasts. Group 2 had no treatment as negative control, group 3 was treated by PRP 10% as positive control group and groups 4-6 were treated by various concentrations of AFM (3.9; 15.625 and 62.5 µg/mL). At the end of the experiment, the proliferation was assessed with MTT assay, furthermore collagen deposition was measured by Sirius red assay. Real Time-PCR (RT-PCR) was performed to quantify Coll I, Coll III and MMP-1 mRNA expression, then to measured COL 1/COL III ratio. Results: UVB induced significant lower viability, upregulated MMP-1 and downregulated COL I and COL III mRNA expressions. Meanwhile AFM treated groups demonstrated higher cell viability with downregulation of MMP-1 and upregulation of COL I and COL III mRNA expressions. The ratio of COL I/ III expression was significantly (p<0.05) lower in the AFM treated groups compared to the UVB group. Among AFM treated groups, administration of 62.5 µg/mL AFM represented the best result. Conclusion: AFM may ameliorate viability of UVB-irradiated human fibroblast culture which associates with downregulating MMP-1, upregulating COL I and Col III, and reducing COL I/III ratio.

Calcitriol Ameliorates Kidney Injury Through Reducing Podocytopathy, Tubular Injury, Inflammation and Fibrosis in 5/6 Subtotal Nephrectomy Model in Rats.

Chronic kidney diseases (CKDs) lead to end-stage renal diseases (ESRD) which are characterized by glomerulosclerosis, tubular injury, anemia, inflammation, and interstitial fibrosis. Vitamin D is known to have renal protective effects. However, its effects relate to low and high doses of Vitamin D in CKD model is still unknown. CKD was performed using 5/6 subtotal nephrectomy procedure in male Sprague Dawley rats (3 months old, 200-300 grams, SN group; n=6), then rats were sacrificed on day 14 after operation. Sham operation was used for control (SO group; n=6). Calcitriol was administered in two doses : 0.01 µg/mL/100 gramsBW/day (SND1 group; n=6) and 0.05 µg/mL/100 gramsBW/day (SND2 group; n=6) intraperitoneally for 14 days. Glomerulosclerosis and tubular injury score were examined using PAS staining, meanwhile, interstitial fibrosis area fraction was assessed with Sirius Red staining. RT-PCR was performed for assessing nephrin, podocin, IL-6, CD68, Collagen-1, and TGF-ß1 mRNA expressions. Immunostaining (IHC) was carried out to observe macrophage (CD68) and myofibroblast (α-SMA). SN demonstrated CKD condition with higher tubular injury, glomerulosclerosis, interstitial fibrosis, and inflammation compared to SO. Calcitriol-treated group (especially SND2) demonstrated significant lower tubular injury, glomerulosclerosis, and interstitial fibrosis compared to SN. SND2 group showed not only significantly lower CD68, IL-6, Collagen-1, and TGF-ß1 mRNA expressions, but also higher mRNA expressions of nephrin and podocin. SND2 group also demonstrated reduction of macrophages infiltration and myofibroblasts expansion based on its histopathological appearance. Vitamin D may have a renoprotective effect on 5/6 subtotal nephrectomy model by attenuating podocytopathy, tubular injury, inflammation and interstitial fibrosis.

Vitamin D Ameliorates Kidney Ischemia Reperfusion Injury via Reduction of Inflammation and Myofibroblast Expansion.

The incidence rate of Acute Kidney Injury (AKI) gets escalated each year. Kidney ischemia/reperfusion injury (IR injury) is the main cause of AKI after major cardiovascular surgery, trauma, or kidney transplantation. Reperfusion is considered essential for ischemic tissue. However, the evidence revealed that reperfusion itself has impact in cellular destruction. Vitamin D is not only known as calcium regulating hormone, but also as renoprotective agent. This study aimed to investigate the effect of vitamin D treatment on kidney IR injury in mice. Kidney IR injury was performed using 30 minutes of bilateral clamping of renal pedicles, then released in male Swiss Webster mice (3 months, 30-40 grams, n=20), which were divided into three groups: sham operation (SO) group, IR injury (IRI) group, and IR injury with 0.25 µg/ kg body weight of vitamin D treatment (IR7+VD). Mice were terminated at day 7 post operation, kidneys were harvested and used for paraffin making, immunostaining and RNA extraction. Tubular injury was quantified based on Periodic Acid-Schiff's (PAS) staining. Immunostaining was done for quantification of macrophage (CD68) and myofibroblast (α-SMA). Reverse Transcriptase PCR (RT-PCR) was done to examine Monocyte Chemoattractant Protein-1 (MCP-1) and Toll-like Receptor 4 (TLR4) mRNA expression. Kidney IR injury induced significant increase of tubular injury, which was associated with higher myofibroblast and macrophage number. Meanwhile, Vitamin D treatment significantly reduced tubular, myofibroblast and macrophage number. RTPCR revealed reduction of TLR4 and MCP-1 mRNA expressions after Vitamin D treatment (p<0.05 vs IR group). Vitamin D ameliorates kidney IR injury through reducing inflammation and myofibroblast formation.

Chlorogenic acid ameliorates memory loss and hippocampal cell death after transient global ischemia.

Chlorogenic acid (CGA) is known to have antioxidant potentials, yet the effect of CGA on brain ischemia has not been sufficiently understood. Brain ischemia such as transient global ischemia disrupts many areas of the brain of rats, including the hippocampus. Male Wistar rats were randomly assigned into five groups, that is, sham-operated (SO), bilateral common carotid occlusion (BCCO), and BCCO+ 15, 30, and 60 mg/kg bw CGA groups (CGA15, CGA30, and CGA60, respectively). Brain ischemia was induced in Wistar rats with BCCO for 20 min followed by intraperitoneal injection of CGA. The rats were examined for the spatial memory in a Morris water maze test on the 3rd day and were euthanized on the 10th day after BCCO. The total number of pyramidal cells was estimated, and the mRNA expressions of Bcl2, Bax, caspase-3, SOD2, SOD1, GPx, ET-1, eNOS, CD31, and VEGF-A were measured. The BCCO group spent less time and distance in the target quadrant than any other group in the spatial memory retention test. The CA1 pyramidal cell numbers in the BCCO and CGA15 groups were lower than in the CGA30 and CGA60 groups. The mRNA expressions of Bcl2, SOD2, and CD31 in the BCCO group were lower than in the CGA15, CGA30, and CGA60 groups. The ET-1 expression was higher in the BCCO and CGA15 groups than in the SO, CGA30, and CGA60 groups. CGA improves the spatial memory and prevents the CA1 pyramidal cell death after BCCO by increasing Bcl2, SOD2, and CD31 expressions and decreasing ET-1 expression.

Ethanol Extract of Centella asiatica (Gotu Kola) Attenuates Tubular Injury Through Inhibition of Inflammatory Cytokines and Enhancement of Anti-Fibrotic Factor in Mice with 5/6 Subtotal Nephrectomy.

BACKGROUND: Chronic kidney disease (CKD) leads to inflammation, fibrosis and destruction of the renal architecture. Centella asiatica (CeA) is an herbaceous plant with anti-inflammatory effects. We aimed to elucidate the effect of CeA on inflammation, fibrosis, vascular remodelling and antifibrotic substances in a 5/6 subtotal nephrectomy (SN) model in mice. METHODS: Mice were divided into three groups: sham operation (SO, n = 6), 5/6 SN for seven days (SN7, n = 7) and SN7 with oral CeA treatment (SN7-CeA, n = 7). At day 7, mice were euthanised, kidneys were harvested and stained with periodic-acid Schiff (for tubular injury and glomerulosclerosis) and sirius red (for fibrosis and vascular remodeling) staining. mRNA expression of prepro-endothelin-1, nephrin, E-cadherin, bone morphogenic protein-7 (BMP-7), toll-like receptor 4 (TLR4), tumour necrosis factor-α (TNFα) and hepatocyte growth factor (HGF) were quantified using reverse transcriptase-PCR. RESULTS: SN group demonstrated significant higher interstitial fibrosis, vascular remodeling, tubular injury and glomerulosclerosis (P < 0.01) compared to SO group. Meanwhile, in SN7-CeA demonstrated attenuation of vascular remodeling as shown by significant higher lumen area with lower Wall/Lumen area ratio compared to SN7. RT-PCR analysis showed up-regulation of nephrin, BMP-7 and E-cadherin mRNA expression (P < 0.05) and down-regulation of ppET-1 in SN7-CeA group compared to SN7 group (P < 0.05). CONCLUSION: CeA may ameliorate renal injury in the SN model in mice.