UC-MSCs Secretome Induces Proliferation of CD4+ T Cells, CD8+ T Cells, NK Cells, and Increases sPD-1 Levels in Severe COVID-19’s Whole Blood

Background: Clinical features of severe coronavirus disease 2019 (COVID-19) predominantly include respiratory symptoms and exacerbated multi-organ complications, especially in patients with comorbidities. Cellular immunity, including lymphocytes, is a critical factor in combating SARS-CoV-2 infection. However, immune dysregulation occurs in severe COVID-19 patients, characterized by cytokine storm and lymphopenia. The effectiveness of mesenchymal stem cell (MSC) therapies for COVID-19 is being assessed. The secretome released by MSC functions similarly to the cells themselves as an immunomodulator, offering potential advantages in terms of safety and cost-effectiveness. This study was conducted to assess the effect of umbilical cord MSC-derived (UC-MSC) secretome treatment on lymphocyte count and soluble programmed cell death-1 (sPD-1) levels in severe COVID-19 patient's whole blood.Materials and methods: Twelve whole blood samples from healthy individuals and severe COVID-19 patients were analyzed for lymphocyte count and functional activation using flow cytometry, along with sPD-1 level measurement in pre-treatment and post-secretome conditions.Results: The lymphocyte count in severe COVID-19 patients was significantly decreased, particularly for T cells and NK cells, indicating lymphopenia. Following secretome treatment, CD4+ T cell counts significantly increased compared to pre-treatment, although this change was not significant in the negative control group. Additionally, there was a minimal reduction in B cell count and an increase in sPD-1 levels. Elevated sPD-1 may alleviate T cell exhaustion by interfering with PD-1 binding to programmed death-ligand 1 (PD-L1).Conclusion: Administration of UC-MSC secretome to the whole blood of severe COVID-19 patients suggested immune improvement, with significant increases in CD4+ T cell counts, enhanced B cell survival, and elevated sPD-1 levels. Keywords: COVID-19, cellular immunity, lymphocytes, secretome, MS

The μDrop Method Enhances Melanin Content Measurement in the in vitro Melanogenesis Model Using B16F10 Cell Line

Background: The B16F10 cell line is a cell frequently used in melanin content assays. However, reports on cell models using B16F10 are limited, particularly as the robust model cell in the Indonesian cosmetics industry. We found measuring melanin content using microplate spectrophotometry to be challenging, so this research was conducted to develop a method using μDrop spectrophotometry.Materials and methods: In this in vitro study, the B16F10 melanoma cell line was cultured in Roswell Park Memorial Institute (RPMI) medium containing 5% fetal bovine serum (FBS). The cells were categorized into control, stimulated, and treated groups. Melanogenesis stimulation was achieved using 1μM α-melanocyte-stimulating hormone (α-MSH), while inhibition using 800 μg/ml kojic acid. After treatment, the cells were incubated for 48 hours. Their melanin content was then measured using an ELISA reader with a μDrop method and compared with the microplate method. Statistical analysis used a one-way ANOVA test with Turkey’s Post Hoc analysis.Results: The μDrop method increased the melanin signal into the linear range of machine readings, while the signals from the microplate method fell far below this range. The B16F10 melanoma cell lines stimulated by α-MSH exhibited increased melanin production compared with the control group, while kojic acid treatment significantly reduced (p<0.05) melanin content in the stimulated group.Conclusion: The μDrop method significantly outperformed the microplate method in measuring melanin content within melanogenesis cell models, offering enhanced accuracy and particularly excelling at quantifying low content of melanin. Keywords: μDrop, microplate, melanin, melanogenesis, B16F10 cell line, RPM

Mesenchymal Stem Cell-Derived Exosomes Enhance FGF-1 and SDF-1 Expression in Rats with Second Degree Burns

Background: Second-degree burns cause extensive damage to the skin and pose significant health challenges, with current treatments facing limitations such as donor skin shortages and complications. Fibroblast growth factor 1 (FGF-1) and stromal-derived growth factor 1 (SDF-1) are critical for tissue repair. Emerging evidence suggests that mesenchymal stem cell-derived exosomes (E-MSCs) are a promising cell-free therapeutic option for enhancing wound healing through the modulation of FGF-1 and SDF-1. This study investigated the effect of E-MSCs on the expression of FGF-1 and SDF-1 genes in rats with second-degree burns.Materials and methods:  This experimental study used a second-degree burn model in Wistar rats, treated with subcutaneous injections of E-MSCs at doses of 100 µL and 200 µL. Gene expression of FGF-1 and SDF-1 was quantified using qRT-PCR. Histological validation confirmed burn severity, and flow cytometry was used to characterize E-MSCs and exosomes.Results: An increase in FGF-1 and SDF-1 expression was observed in exosome-treated groups compared to the NaCL-treated group. The 200 µL E-MSCs-treated group showed the most significant enhancement in both growth factors, with statistically significant differences (p<0.05). These findings underline the efficacy of E-MSCs in modulating critical genes involved in wound healing.Conclusion: E-MSCs significantly upregulate FGF-1 and SDF-1 expression, promoting tissue repair and regeneration in second-degree burn models. This study highlights the potential of E-MSCs as a non-invasive therapeutic approach.  Keywords: exosomes, FGF-1, mesenchymal stem cells, SDF-

The Role of Centella asiatica and Its Main Bioactive Compound, Asiatic Acid in Cardiac Remodeling: A Systematic Review of Animal Studies

Cardiac remodeling is a phenotype of heart failure characterized by a molecular, cellular, and interstitial change in the heart, which manifests in the change of size and function of the heart after specific insults with multiple mechanisms. Centella asiatica (CA) and its main bioactive triterpenoid, asiatic acid (AA), pose antioxidant and anti-inflammatory effects. Still, no adequate clinical trials support the potency of CA and AA as anti-cardiac remodeling. Hence, this systematic review aims to provide an in-depth analysis of CA extract and AA in animal studies' prevention or therapy of cardiac remodeling. The search strategies were conducted based on preferred reporting Items for systematic reviews and meta-analysis (PRISMA) protocol through Pubmed, EMBASE, Scopus, and Web of Science using keywords as follows: “Centella asiatica” OR “Asiatic Acid” AND “Cardiac Remodeling” OR “Cardiac Hypertrophy” OR “Cardiac Fibrosis” along with their synonym. The data collected included hemodynamic parameters based on echocardiography, biomolecular tests such as quantitative polymerase chain reaction (qPCR), Western blotting, or biochemistry procedures. The paper quality was assessed using Systematic Review Centre for Laboratory Animal Experimentation (SYRCLE) risk of bias (RoB). The previous selected study has shown that CA and AA might prevent and cure cardiac remodeling by inhibiting various pathways and protein expressions through AMPKα, NOX2/4, PI3K/Akt/mTOR, p70S6K, YAP/TAZ, and IL-1β, IL-6, and IL-18 cytokines. CA and AA, thus, exhibit cardioprotective effects in the animal model, which need to be confirmed in the clinical trials on humans. Keywords: cardiac remodeling, cardiac hypertrophy, cardiac fibrosis, Centella asiatica, asiatic aci

Resveratrol Protects Caenorhabditis elegans from Ultraviolet B-induced Photoaging via skn-1

Background: Resveratrol (RSV) is a polyphenol with potent antioxidant activity and is abundant in fruits. There has been a lot of scientific evidence regarding the anti-aging effect of RSV. Aging can be induced by UV-B (photoaging) due to the production of reactive oxygen species (ROS) and oxidative stress. This study aimed to test the anti-photoaging activity of RSV on UV-B -induced Caenorhabditis elegans.Materials and methods: C. elegans was cultured at 20˚C in nematode growth medium (NGM) and was subjected to various concentrations of RSV and UV-B. The UV-B light exposure was given on day 0 post-synchronization at a dose of 100 J/m2 using a UV cross-linker. The health span (indicated by pharyngeal pumping rate) and lifespan of worms were observed. The quantification of collagen was performed using a Sircol Collagen assay kit. The mRNA expression level of gcs-1, col-19, hus-1, cep-1, egl-1, and ced-13 was examined by qRT-PCR. Results: UV-B reduced pharyngeal pumping rate, shortened the lifespan, decreased collagen, and increased the expression of apoptosis-related genes (hus-1, cep-1, egl-1, and ced-13). RSV ameliorated these aging phenotypes induced by UV-B. Anti-aging activities of RSV were not observed in the skn-1 loss-of-function strain (VC1772, skn-1(ok2315)), indicating the critical involvement of skn-1 in the mechanism of action of RSV. The activation of skn-1 was shown by elevated skn-1 target gene that play role in glutathione biosynthesis called gcs-1.Conclusion: RSV prevents accelerated aging due to UV-B in C. elegans by enforcing skn-1 signaling pathway and its downstream gcs-1 gene expression.   Keywords: anti-aging, resveratrol, oxidative stress, UV-

Sorghum-Soybean Flour Enteral Formula Reduces Blood Glucose, Cholesterol, Triglycerides, LDL, and Increases HDL and Albumin in Hyperglycemic Rats

Background: Diabetes mellitus prevalence is rising. Liquid feeding in the form of enteral formulas is needed to meet the nutritional needs of patients who cannot consume orally. The development of enteral formulas based on sorghum-soybean flour for diabetes mellitus patients, which has been nutritionally analyzed and adjusted to the requirements of enteral manufacturing, was selected for further in vivo research. This study evaluated the effect of the sorghum-soybean flour formula on fasting blood glucose (FBG), lipid profile levels, and albumin in hyperglycemic rats.Materials and methods: This was a true experimental study with pre–post-test randomized control design. Wistar rats were divided into four groups: negative control group was normal rats and given standard feed only; positive control group was hyperglycemic rats and given standard feed only; treatment (T)1 and T2 groups were hyperglycemic rats given standard feed along with enteral formula at a dose of 4,41 g/day and 5,51 g/day for 28 days. Blood samples were collected to analyze FBG, albumin, low-density lipoprotein (LDL), high-density lipoprotein (HDL), triglycerides, and total cholesterol.Results: There were differences in the levels of FBG, albumin, LDL, HDL, triglycerides, and total cholesterol before and after the intervention in groups T1 and T2. Group T1 showed an 8.12% decrease in FBG, while T2 showed a 29.89% decrease. Triglycerides decreased by 29.22% in T1 and 31.85% in T2; cholesterol decreased by 11.41% in T1 and 13.94% in T2. LDL levels decreased by 29.97% in T1 and 38.44% in T2. Albumin levels increased by 47.90% in T1 and 56.67% in T2. HDL levels increased by 23.94% in T1 and in 35.04% in T2.Conclusion: Administration of an enteral formula based on sorghum-soybean flour can reduce FBG, triglycerides, total cholesterol, and LDL levels, and increase albumin and HDL levels in hyperglycemic rats.Keywords: hyperglycemia, enteral formula, albumin levels, fasting blood glucose, HDL, LDL, total cholesterol, triglyceride

Green Tea Yogurt Supplemented with L. paracasei E1 Microcapsules Increases Erythrocyte Counts and B Cell Development in High-Fat Fructose Diet Mice

Background: Obesity-induced inflammation causes hematopoietic stress, disrupting bone marrow homeostasis. Green tea yogurt supplemented with Lacticaseibacillus paracasei E1 microcapsules (GTYP) is a promising way to overcome obesity due to its high antioxidant and anti-inflammatory activity. However, GTYP effects on blood production, specifically erythrocytes and B cells, remain unexplored.Materials and methods: Male Balb/C mice were fed either a high-fat fructose diet or a normal diet for 12 weeks. Microencapsulation was done by double coating of alginate-chitosan. There were seven groups in this study: normal diet (ND), high-fat fructose diet (HFFD), HFFD with 1.3 mg/kgBW Simvastatin (T1), HFFD with 5 g/kgBW plain yogurt (T2), HFFD with 2.5 g/kgBW GTYP (T3), HFFD with 5 g/kgBW GTYP (T4), and HFFD with 10 g/kgBW GTYP (T5). Erythrocyte counts from the peripheral blood were taken weekly. After 28 days of treatment, mice were sacrificed, bone marrow (BM) and lymphocytes were isolated. The cells of Ter119+, Ter119+CD59+, and B220+SDF-1+ were measured using flow cytometry.Results: HFFD not only reduces the peripheral erythrocyte count (2.15×109 cell/mL) but also affects the hematopoietic system, depleting Ter119+ (11.76%), TER119+CD59+ (0.050%), and B220+SDF-1+ (0.465%). Mice receiving 5 g/KgBW GTYP improved erythrocyte count (9.95×109 cells/mL). The parameters of erythrocyte and B cell development showed more remarkable improvement with GTYP treatment than simvastatin and plain yogurt (p<0.05). Molecular docking also indicated a great inhibitory effect of EGCG (-7.7) for the CXCR4 receptor.Conclusion: GTYP can potentially increase erythrocyte count and B cell development, particularly under obese conditions.   Keywords: anti-obesity, B lymphopoiesis, erythrocyte count, green tea yogurt, probiotic

n-Hexane Fraction of Cucumis melo L. Cultivar Gama Melon Parfum: An in vitro Study in MCF7 and T47D Cells Line

Background: Cucumis melo a melon species, typically has a sweet taste. Some cultivars are known for their distinctive bitter flesh due to its higher levels of cucurbitacin. Cucurbitacin is semipolar compound that has anticancer properties. However, the anticancer effects of cucurbitacin from gama melon parfum (GMP) have not been widely studied. The use of n-Hexane as a non-polar solvent in GMP melon fractionation is to dissolve the non-polar parts of the plant. However, Cucurbitacin was found in the n-hexane fraction of Cucurbita pepo L. Therefore, this study will investigate the presence of Cucurbitacin in the n-Hexane fraction and its effects on breast cancer cells T47D and MCF7.Materials and methods: Dry simplicia of GMP melon fruit were macerated using methanol and fractionated using n-hexane. The presence of cucurbitacin was detected using the high-performance liquid chromatography (HPLC) method. Cell cytotoxicity tests were assessed using the MTT assay, with concentrations of 7.8125, 15.625, 31.25, 62.5, and 125 µg/mLResults: Cucurbitacin compounds were detected in the n-hexane fraction at a concentration of 7.6 µg/mL per 10 mg of n-hexane fraction. MCF7 cell viability was lower than that of T47D cells across all concentrations tested. MCF7 cell viability was below 50% at a concentration of 62.5 µg/mL. In contrast, T47D cell viability remained at 100% even at the highest concentration of 125 µg/mL. The IC50 value of MCF7 cells was 43.5 µg/mL.Conclusion: The cucurbitacin content in the n-Hexane fraction was 7.6 µg/mL per 10mg fraction. At this concentration, it moderately inhibits the proliferation of MCF7 cells.Keywords: gama melon parfum, cucurbitacin, HPLC, T47D, MCF

Hypoxia-Induced Mesenchymal Stem Cell Exosomes Modulate Protein Kinase A and VEGFR Expression in Ultraviolet B-Induced Hyperpigmentation in Mice

Background: Hyperpigmentation is often exacerbated by ultraviolet-B (UVB) exposure through oxidative stress and activation of pathways like mitogen-activated protein kinase (MAPK) and vascular endothelial growth factor receptor (VEGFR). Current treatments carry risks and necessitate safer alternatives. This study investigated the therapeutic potential of hypoxia-induced mesenchymal stem cell (MSC) exosomes in reducing protein kinase-A (PKA) and VEGFR expression in UVB-induced hyperpigmentation.Materials and methods: A post-test-only control group design was used with 30 male C57BL/6 mice divided into five groups: Healthy group, 0,9% NaCl-treated group, retinol-treated group, and two treatment groups (200 µL Exosomes-treated group and 300 µL Exosomes-treated group. UVB-induced hyperpigmentation was established with 180 mJ/cm² exposures over two weeks. Treatment was administered via subcutaneous injections for seven days. PKA and VEGFR mRNA levels were analyzed using qRT-PCR.Results: PKA expression was significantly lower in the 200 µL Exosomes-treated group (0.34±0.05) and 300 µL Exosomes-treated group (0.21±0.04) groups compared with the 0,9% NaCl-treated group (1.12±0.08) (p<0.001). VEGFR expression similarly decreased in 200 µL Exosomes-treated group (0.32±0.05) and 300 µL Exosomes-treated group (0.18±0.04) versus the 0,9% NaCl-treated group (1.48±0.09) (p<0.001). Both exosome doses achieved reductions comparable to baseline levels observed in the Healthy group.Conclusion: Hypoxia-induced MSC exosomes reduced PKA and VEGFR expression in UVB-induced hyperpigmentation, with the 300 µL dose showing greater efficacy. These findings suggested exosome therapy as a promising alternative for hyperpigmentation treatment. Keywords: hyperpigmentation, MSC, PKA, VEGFR, melani

Slow 0.9% NaCl Bolus Administration Reduces ANP, MMP-2, and Syndecan-1 Shedding in Septic Shock Rabbit Models

Background: The optimal rate for fluid bolus administration in septic shock remains a critical and unresolved question. Rapid bolus administration is commonly practiced but has been linked to elevated levels of atrial natriuretic peptide (ANP), matrix metalloproteinase-2 (MMP-2), and syndecan-1 shedding, potentially exacerbating endothelial glycocalyx damage and increasing vascular permeability. However, the physiological and clinical implications of slower bolus rates have not been thoroughly investigated. This study was conducted to identify safer fluid management practices and improve patient outcomes in septic shock.Materials and methods: A randomized post-test-only control group design was employed, involving 36 male New Zealand rabbits with lipopolysaccharide-induced septic shock. The treatment group received 0.9% NaCl boluses (20 mL/kg body weight) over 20 minutes per bolus (slow bolus), while the control group received the same volume over 5 minutes per bolus (rapid bolus). ANP, MMP-2, and syndecan-1 levels were measured using ELISA 10-15 minutes post-intervention.Results: The median ANP levels in the treatment group (92.86 ng/mL) were significantly lower (p<0.05) than those in the control group (367.32 ng/mL). The mean MMP-2 levels in the treatment group (10.26 ng/dL) were lower than those in the control group (11.43 ng/dL). The median levels of syndecan-1 were also lower in the treatment group (4.31 ng/mL) compared to the control group (5.94 ng/mL).Conclusion: Slow fluid boluses appear to mitigate endothelial damage by reducing ANP, MMP-2, and syndecan-1 shedding. These findings suggest that slower infusion rates may offer a protective advantage in fluid resuscitation, paving the way for updated clinical guidelines.Keywords: fluid bolus, ANP, MMP-2, syndecan-