Canthin-6-One Inhibits Developmental and Tumour-Associated Angiogenesis in Zebrafish.

Tumour-associated angiogenesis play key roles in tumour growth and cancer metastasis. Consequently, several anti-angiogenic drugs such as sunitinib and axitinib have been approved for use as anti-cancer therapies. However, the majority of these drugs target the vascular endothelial growth factor A (VEGFA)/VEGF receptor 2 (VEGFR2) pathway and have shown mixed outcome, largely due to development of resistances and increased tumour aggressiveness. In this study, we used the zebrafish model to screen for novel anti-angiogenic molecules from a library of compounds derived from natural products. From this, we identified canthin-6-one, an indole alkaloid, which inhibited zebrafish intersegmental vessel (ISV) and sub-intestinal vessel development. Further characterisation revealed that treatment of canthin-6-one reduced ISV endothelial cell number and inhibited proliferation of human umbilical vein endothelial cells (HUVECs), suggesting that canthin-6-one inhibits endothelial cell proliferation. Of note, canthin-6-one did not inhibit VEGFA-induced phosphorylation of VEGFR2 in HUVECs and downstream phosphorylation of extracellular signal-regulated kinase (Erk) in leading ISV endothelial cells in zebrafish, suggesting that canthin-6-one inhibits angiogenesis independent of the VEGFA/VEGFR2 pathway. Importantly, we found that canthin-6-one impairs tumour-associated angiogenesis in a zebrafish B16F10 melanoma cell xenograft model and synergises with VEGFR inhibitor sunitinib malate to inhibit developmental angiogenesis. In summary, we showed that canthin-6-one exhibits anti-angiogenic properties in both developmental and pathological contexts in zebrafish, independent of the VEGFA/VEGFR2 pathway and demonstrate that canthin-6-one may hold value for further development as a novel anti-angiogenic drug.

Mechanistic insight of α-mangostin encapsulation in 2-hydroxypropyl-ß-cyclodextrin for solubility enhancement.

α-Mangostin is the most abundant compound contained in the mangostin (Garcinia mangostana L.) plant which have been developed and proven to have many promising pharmacological effects. However, the low water solubility of α-mangostin causes limitations in its development in clinical purpose. To increase the solubility of a compound, a method currently being developed is to make drug inclusion complexes using cyclodextrins. This research aimed to use in silico techniques namely molecular docking study and molecular dynamics simulation to explore the molecular mechanism and stability of the encapsulation of α-mangostin using cyclodextrins. Two types of cyclodextrins were used including ß-cyclodextrin and 2-hydroxypropyl-ß-cyclodextrin docked against α-mangostin. From the molecular docking results, it shows that the α-mangostin complex with 2-hydroxypropyl-ß-cyclodextrin provides the lowest binding energy value of -7.99 Kcal/mol compared to ß-cyclodextrin value of -6.14 Kcal/mol. The α-mangostin complex with 2-hydroxypropyl-ß-cyclodextrin also showed good stability based on molecular dynamics simulation during 100 ns. From molecular motion, RDF, Rg, SASA, density, total energy analyzes, this complex shows increased solubility in water and provided good stability. This indicates that the encapsulation of α-mangostin with 2-hydroxypropyl-ß-cyclodextrin can increase the solubility of the α-mangostin.Communicated by Ramaswamy H. Sarma.

Human ACE2 orthologous peptide sequences show better binding affinity to SARS-CoV-2 RBD domain: Implications for drug design.

Computational methods coupled with experimental validation play a critical role in the identification of novel inhibitory peptides that interact with viral antigenic determinants. The interaction between the receptor binding domain (RBD) of SARS-CoV-2 spike protein and the helical peptide of human angiotensin-converting enzyme-2 (ACE2) is a necessity for the initiation of viral infection. Herein, natural orthologs of human ACE2 helical peptide were evaluated for competitive inhibitory binding to the viral RBD by use of a computational approach, which was experimentally validated. A total of 624 natural ACE2 orthologous 32-amino acid long peptides were identified through a similarity search. Molecular docking was used to virtually screen and rank the peptides based on binding affinity metrics, benchmarked against human ACE2 peptide docked to the RBD. Molecular dynamics (MD) simulations were done for the human reference and the Nipponia nippon peptide as it exhibited the highest binding affinity (Gibbs free energy; -14 kcal/mol) predicted from the docking results. The MD simulation confirmed the stability of the assessed peptide in the complex (-12.3 kcal/mol). The top three docked-peptides (from Chitinophaga sancti, Nipponia nippon, and Mus musculus) and the human reference were experimentally validated by use of surface plasmon resonance technology. The human reference exhibited the weakest binding affinity (Kd of 318-441 pM) among the peptides tested, in agreement with the docking prediction, while the peptide from Nipponia nippon was the best, with 267-538-fold higher affinity than the reference. The validated peptides merit further investigation. This work showcases that the approach herein can aid in the identification of inhibitory biosimilar peptides for other viruses.

Antioxidant, Wound Healing Potential and In Silico Assessment of Naringin, Eicosane and Octacosane.

1. Diabetic chronic wounds, mainly foot ulcers, constitute one of the most common complications of poorly managed diabetes mellitus. The most typical reasons are insufficient glycemic management, latent neuropathy, peripheral vascular disease, and neglected foot care. In addition, it is a common cause of foot osteomyelitis and amputation of the lower extremities. Patients are admitted in larger numbers attributable to chronic wounds compared to any other diabetic disease. In the United States, diabetes is currently the most common cause of non-traumatic amputations. Approximately five percent of diabetics develop foot ulcers, and one percent require amputation. Therefore, it is necessary to identify sources of lead with wound-healing properties. Redox imbalance due to excessive oxidative stress is one of the causes for the development of diabetic wounds. Antioxidants have been shown to decrease the progression of diabetic neuropathy by scavenging ROS, regenerating endogenous and exogenous antioxidants, and reversing redox imbalance. Matrix metalloproteinases (MMPs) play vital roles in numerous phases of the wound healing process. Antioxidant and fibroblast cell migration activity of Marantodes pumilum (MP) crude extract has previously been reported. Through their antioxidant, epithelialization, collagen synthesis, and fibroblast migration activities, the authors hypothesise that naringin, eicosane and octacosane identified in the MP extract may have wound-healing properties. 2. The present study aims to identify the bioactive components present in the dichloromethane (DCM) extract of M. pumilum and evaluate their antioxidant and wound healing activity. Bioactive components were identified using LCMS, HPTLC and GCMS. Excision wound on STZ-induced diabetic rat model, human dermal fibroblast (HDF) cell line and colorimetric antioxidant assays were used to evaluate wound healing and antioxidant activities, respectively. Molecular docking and pkCMS software would be utilised to predict binding energy and affinity, as well as ADME parameters. 3. Naringin (NAR), eicosane (EIC), and octacosane (OCT) present in MP displayed antioxidant action and wound excision closure. Histological examination HDF cell line demonstrates epithelialization, collagen production, fibroblast migration, polymorphonuclear leukocyte migration (PNML), and fibroblast movement. The results of molecular docking indicate a substantial attraction and contact between MMPs. pkCMS prediction indicates inadequate blood-brain barrier permeability, low toxicity, and absence of hepatotoxicity. 4. Wound healing properties of (NEO) naringin, eicosane and octacosane may be the result of their antioxidant properties and possible interactions with MMP.

Role of Probiotics in the Management of COVID-19: A Computational Perspective.

Coronavirus disease 2019 (COVID-19) was declared a pandemic at the beginning of 2020, causing millions of deaths worldwide. Millions of vaccine doses have been administered worldwide; however, outbreaks continue. Probiotics are known to restore a stable gut microbiota by regulating innate and adaptive immunity within the gut, demonstrating the possibility that they may be used to combat COVID-19 because of several pieces of evidence suggesting that COVID-19 has an adverse impact on gut microbiota dysbiosis. Thus, probiotics and their metabolites with known antiviral properties may be used as an adjunctive treatment to combat COVID-19. Several clinical trials have revealed the efficacy of probiotics and their metabolites in treating patients with SARS-CoV-2. However, its molecular mechanism has not been unraveled. The availability of abundant data resources and computational methods has significantly changed research finding molecular insights between probiotics and COVID-19. This review highlights computational approaches involving microbiome-based approaches and ensemble-driven docking approaches, as well as a case study proving the effects of probiotic metabolites on SARS-CoV-2.

Potential Adjuvant Therapeutic Effect of Lactobacillus plantarum Probio-88 Postbiotics against SARS-COV-2.

In response to the ongoing COVID-19 pandemic, the global effort to develop high efficacy countermeasures to control the infection are being conducted at full swing. While the efficacy of vaccines and coronavirus drugs are being tested, the microbiome approach represents an alternative pathophysiology-based approach to prevent the severity of the infection. In the current study, we evaluated the action of a novel probiotic Lactobacillus plantarum Probio-88 against SARS-COV-2 replication and immune regulation using an in vitro and in silico study. The results showed that extract from this strain (P88-CFS) significantly inhibited the replication of SARS-COV-2 and the production of reactive oxygen species (ROS) levels. Furthermore, compared with infected cells, P88-CFS treated cells showed a significant reduction in inflammatory markers such as IFN-α, IFN-ß, and IL-6. Using an in silico molecular docking approach, it was postulated that the antiviral activity of L. plantarum Probio-88 was derived from plantaricin E (PlnE) and F (PlnF). The high binding affinity and formation of hydrogen bonding indicated that the association of PlnE and PlnF on SARS-COV-2 helicase might serve as a blocker by preventing the binding of ss-RNA during the replication of the virus. In conclusion, our study substantiated that P88-CFS could be used as an integrative therapeutic approach along with vaccine to contain the spread of the highly infectious pathogen and possibly its variants.

Drug Discovery of Spinal Muscular Atrophy (SMA) from the Computational Perspective: A Comprehensive Review.

Spinal muscular atrophy (SMA), one of the leading inherited causes of child mortality, is a rare neuromuscular disease arising from loss-of-function mutations of the survival motor neuron 1 (SMN1) gene, which encodes the SMN protein. When lacking the SMN protein in neurons, patients suffer from muscle weakness and atrophy, and in the severe cases, respiratory failure and death. Several therapeutic approaches show promise with human testing and three medications have been approved by the U.S. Food and Drug Administration (FDA) to date. Despite the shown promise of these approved therapies, there are some crucial limitations, one of the most important being the cost. The FDA-approved drugs are high-priced and are shortlisted among the most expensive treatments in the world. The price is still far beyond affordable and may serve as a burden for patients. The blooming of the biomedical data and advancement of computational approaches have opened new possibilities for SMA therapeutic development. This article highlights the present status of computationally aided approaches, including in silico drug repurposing, network driven drug discovery as well as artificial intelligence (AI)-assisted drug discovery, and discusses the future prospects.

Lactobacillus Strains Alleviated Hyperlipidemia and Liver Steatosis in Aging Rats via Activation of AMPK.

In this study, we hypothesized that different strains of Lactobacillus can alleviate hyperlipidemia and liver steatosis via activation of 5' adenosine monophosphate-activated protein kinase (AMPK), an enzyme that is involved in cellular energy homeostasis, in aged rats. Male rats were fed with a high-fat diet (HFD) and injected with D-galactose daily over 12 weeks to induce aging. Treatments included (n = 6) (i) normal diet (ND), (ii) HFD, (iii) HFD-statin (lovastatin 2 mg/kg/day), (iv) HFD-Lactobacillus fermentum DR9 (10 log CFU/day), (v) HFD-Lactobacillus plantarum DR7 (10 log CFU/day), and (vi) HFD-Lactobacillus reuteri 8513d (10 log CFU/day). Rats administered with statin, DR9, and 8513d reduced serum total cholesterol levels after eight weeks (p < 0.05), while the administration of DR7 reduced serum triglycerides level after 12 weeks (p < 0.05) as compared to the HFD control. A more prominent effect was observed from the administration of DR7, where positive effects were observed, ranging from hepatic gene expressions to liver histology as compared to the control (p < 0.05); downregulation of hepatic lipid synthesis and ß-oxidation gene stearoyl-CoA desaturase 1 (SCD1), upregulation of hepatic sterol excretion genes of ATP-binding cassette subfamily G member 5 and 8 (ABCG5 and ABCG8), lesser degree of liver steatosis, and upregulation of hepatic energy metabolisms genes AMPKα1 and AMPKα2. Taken altogether, this study illustrated that the administration of selected Lactobacillus strains led to improved lipid profiles via activation of energy and lipid metabolisms, suggesting the potentials of Lactobacillus as a promising natural intervention for alleviation of cardiovascular and liver diseases.

Molecular evolutionary and 3D protein structural analyses of Lactobacillus fermentum elongation factor Tu, a novel brain health promoting factor.

The role of microbiota in gut-brain communication has led to the development of probiotics promoting brain health. Here we report a genomic study of a Lactobacillus fermentum PS150 and its patented bioactive protein, elongation factor Tu (EF-Tu), which is associated with cognitive improvement in rats. The L. fermentum PS150 circular chromosome is 2,238,401 bp and it consists of 2281 genes. Chromosome comparisons with other L. fermentum strains highlighted a cluster of glycosyltransferases as potential candidate probiotic factors besides EF-Tu. Molecular evolutionary analyses on EF-Tu genes (tuf) in 235 bacteria species revealed one to three copies of the gene per genome. Seven tuf pseudogenes were found and three species only possessed pseudogenes, which is an unprecedented finding. Protein variability analysis of EF-Tu showed five highly variable residues (40 K, 41G, 42 L, 44 K, and 46E) on the protein surface, which warrant further investigation regarding their potential roles as binding sites.

Allantoin, a Potential Metabolite That Promotes AMPK Phosphorylation and Suppresses Cholesterol Biosynthesis Via the Mevalonate Pathway and Bloch Pathway.

This study aimed to evaluate the effect of probiotic administration on obese and ageing models. Sprague Dawley rats were subjected to high-fat diet (HFD) and injected with D-galactose to induce premature ageing. Upon 12 weeks of treatment, the faecal samples were collected and subjected to gas chromatography-mass spectrophotometry (GC-MS) analysis for metabolite detection. The sparse partial least squares discriminant analysis (sPLS-DA) showed a distinct clustering pattern of metabolite profile in the aged and obese rats administered with probiotics Lactobacillus plantarum DR7 and L. reuteri 8513d, particularly with a significantly higher concentration of allantoin. Molecular docking simulation showed that allantoin promoted the phosphorylation (activation) of adenosine monophosphate-activated kinase (AMPK) by lowering the substrate free energy of binding (FEB) and induced the formation of an additional hydrogen bond between Val184 and the substrate AMP. Allantoin also suppressed cholesterol biosynthesis by either inducing enzyme inhibition, occupying or blocking the putative binding site to result in non-spontaneous substrate binding, as in the cases of 3-hydroxy-methylglutaryl-coA reductase (HMGCR), mevalonate kinase (MVK) and lanosterol demethylase (LDM) where positive FEBs were reported. These results demonstrated the potential of allantoin to alleviate age-related hypercholesterolaemia by upregulating AMPK and downregulating cholesterol biosynthesis via the mevalonate pathway and Bloch pathway.

Effects of Potential Probiotic Strains on the Fecal Microbiota and Metabolites of D-Galactose-Induced Aging Rats Fed with High-Fat Diet.

Both aging and diet play an important role in influencing the gut ecosystem. Using premature senescent rats induced by D-galactose and fed with high-fat diet, this study aims to investigate the effects of different potential probiotic strains on the dynamic changes of fecal microbiome and metabolites. In this study, male Sprague-Dawley rats were fed with high-fat diet and injected with D-galactose for 12 weeks to induce aging. The effect of Lactobacillus plantarum DR7, L. fermentum DR9, and L. reuteri 8513d administration on the fecal microbiota profile, short-chain fatty acids, and water-soluble compounds were analyzed. It was found that the administration of the selected strains altered the gut microbiota diversity and composition, even at the phylum level. The fecal short-chain fatty acid content was also higher in groups that were administered with the potential probiotic strains. Analysis of the fecal water-soluble metabolites revealed that administration of L. plantarum DR7 and L. reuteri 8513d led to higher fecal content of compounds related to amino acid metabolism such as tryptophan, leucine, tyrosine, cysteine, methionine, valine, and lysine; while administration of L. fermentum DR9 led to higher prevalence of compounds related to carbohydrate metabolism such as erythritol, xylitol, and arabitol. In conclusion, it was observed that different strains of lactobacilli can cause difference alteration in the gut microbiota and the metabolites, suggesting the urgency to explore the specific metabolic impact of specific strains on the host.

Lactobacillus plantarum USM8613 Aids in Wound Healing and Suppresses Staphylococcus aureus Infection at Wound Sites.

This study aimed to elucidate the targets and mechanisms of anti-staphylococcal effects from bioactive metabolites produced by lactic acid bacteria. We aimed to better understand the safety and efficacy of these bioactive metabolites in in vivo systems, typically at topical sites. The cell-free supernatant and protein-rich fraction from Lactobacillus plantarum USM8613 inhibited staphyloxanthin biosynthesis, reduced (p < 0.05) the cell number of Staphylococcus aureus by 106 CFU/mL and reduced biofilm thickness by 55% in S. aureus-infected porcine skins. Genome-wide analysis and gene expression analysis illustrated the production of several plantaricins, especially the plantaricins EF and JK that enhanced the anti-staphylococcal effects of L. plantarum USM8613. In vivo data using rats showed that the protein-rich fraction from L. plantarum USM8613 exerted wound healing properties via direct inhibition of S. aureus and promoted innate immunity, in which the expression of ß-defensin was significantly (p < 0.05) upregulated by 3.8-fold. The protein fraction from L. plantarum USM8613 also significantly enhanced (p < 0.05) the production of cytokines and chemokines through various stages of wound recovery. Using ∆atl S. aureus, the protein-rich fraction from L. plantarum USM8613 exerted inhibitory activity via targeting the atl gene in S. aureus. Taken altogether, our present study illustrates the potential of L. plantarum USM8613 in aiding wound healing, suppressing of S. aureus infection at wound sites and promoting host innate immunity.

Extracellular transglycosylase and glyceraldehyde-3-phosphate dehydrogenase attributed to the anti-staphylococcal activity of Lactobacillus plantarum USM8613.

Increasing levels of antibiotic resistance in pathogens, including Staphylococcus aureus, remains a serious problem for public health, leading to the need for better alternative antimicrobial strategies. The antimicrobial proteins produced by Lactobacillus plantarum USM8613 attributed to its anti-staphylococcal activity were identified as extracellular transglycosylase and glyceraldehyde-3-phosphate dehydrogenase (GADPH), both with different mechanisms of action. Extracellular transglycosylase, which contains a LysM domain, exerts a cell wall-mediated killing mechanism, while GADPH penetrates into S. aureus cells and subsequently induces the overexpression of autolysis regulators, resulting in S. aureus autolysis. Both extracellular transglycosylase and GADPH exert anti-inflammatory effects in S. aureus-infected HaCaT cells by reducing the expression and production of TLR-2, hBDs and various pro-inflammatory cytokines (IL-1α, IL-1ß, IL-6, TNF-α, and IL-8). Taken together, extracellular transglycosylase and GADPH produced by L. plantarum USM8613 could potentially be applied as an alternative therapeutic agent to treat S. aureus skin infections and promote skin health.

Lactobacillus plantarum DR7 improved upper respiratory tract infections via enhancing immune and inflammatory parameters: A randomized, double-blind, placebo-controlled study.

The aims of this study were to investigate the effects of Lactobacillus plantarum DR7 isolated from bovine milk against upper respiratory tract infections (URTI) and elucidate the possible mechanisms underlying immunomodulatory properties. The DR7 strain (9 log cfu/d) was administered for 12 wk in a randomized, double-blind, and placebo-controlled human study involving 109 adults (DR7, n = 56; placebo, n = 53). Subjects were assessed for health conditions monthly via questionnaires, and blood samples were evaluated for cytokine concentrations, peroxidation and oxidative stress, and gene expression in T cells and natural killer (NK) cells. The administration of DR7 reduced the duration of nasal symptoms (mean difference 5.09 d; 95% CI: 0.42-9.75) and the frequency of URTI (mean difference 0.32; 95% CI: 0.01-0.63) after 12 and 4 wk, respectively, compared with the placebo. The DR7 treatment suppressed plasma proinflammatory cytokines (IFN-γ, TNF-α) in middle-aged adults (30 to 60 yr old), while enhancing anti-inflammatory cytokines (IL-4, IL-10) in young adults (<30 yr old), accompanied by reduced plasma peroxidation and oxidative stress levels compared with the placebo. Young adults who received DR7 showed higher expression of plasma CD44 and CD117 by 4.50- and 2.22-fold, respectively, compared with the placebo. Meanwhile, middle-aged adults showed lower expression of plasma CD4 and CD8 by 11.26- and 1.80-fold, respectively, compared with the placebo, indicating less T-cell activation. In contrast, both young and middle-aged adults who received DR7 showed enhanced presence of nonresting and mature NK cells compared with those who received the placebo. We postulate that DR7 alleviated the symptoms of URTI by improving inflammatory parameters and enhancing immunomodulatory properties.

Applications of Ensemble Docking in Potential Inhibitor Screening for Mycobacterium tuberculosis Isocitrate Lyase Using a Local Plant Database.

Isocitrate lyase (ICL) is a persistent factor for the survival of dormant stage Mycobacterium tuberculosis (MTB), thus a potential drug target for tuberculosis treatment. In this work, ensemble docking approach was used to screen for potential inhibitors of ICL. The ensemble conformations of ICL active site were obtained from molecular dynamics simulation on three dimer form systems, namely the apo ICL, ICL in complex with metabolites (glyoxylate and succinate), and ICL in complex with substrate (isocitrate). Together with the ensemble conformations and the X-ray crystal structures, 22 structures were used for the screening against Malaysian Natural Compound Database (NADI). The top 10 compounds for each ensemble conformation were selected. The number of compounds was then further narrowed down to 22 compounds that were within the Lipinski's Rule of Five for drug-likeliness and were also docked into more than one ensemble conformation. Theses 22 compounds were furthered evaluate using whole cell assay. Some compounds were not commercially available; therefore, plant crude extracts were used for the whole cell assay. Compared to itaconate (the known inhibitor of ICL), crude extracts from Manilkara zapota, Morinda citrifolia, Vitex negundo, and Momordica charantia showed some inhibition activity. The MIC/MBC value were 12.5/25, 12.5/25, 0.78/1.6, and 0.39/1.6 mg/mL, respectively. This work could serve as a preliminary study in order to narrow the scope for high throughput screening in the future.

Thioguanine-based DENV-2 NS2B/NS3 protease inhibitors: Virtual screening, synthesis, biological evaluation and molecular modelling.

Dengue virus Type 2 (DENV-2) is predominant serotype causing major dengue epidemics. There are a number of studies carried out to find its effective antiviral, however to date, there is still no molecule either from peptide or small molecules released as a drug. The present study aims to identify small molecules inhibitor from National Cancer Institute database through virtual screening. One of the hits, D0713 (IC50 = 62 µM) bearing thioguanine scaffold was derivatised into 21 compounds and evaluated for DENV-2 NS2B/NS3 protease inhibitory activity. Compounds 18 and 21 demonstrated the most potent activity with IC50 of 0.38 µM and 16 µM, respectively. Molecular dynamics and MM/PBSA free energy of binding calculation were conducted to study the interaction mechanism of these compounds with the protease. The free energy of binding of 18 calculated by MM/PBSA is -16.10 kcal/mol compared to the known inhibitor, panduratin A (-11.27 kcal/mol), which corroborates well with the experimental observation. Results from molecular dynamics simulations also showed that both 18 and 21 bind in the active site and stabilised by the formation of hydrogen bonds with Asn174.

Probiotic Lactobacillus plantarum P8 alleviated stress and anxiety while enhancing memory and cognition in stressed adults: A randomised, double-blind, placebo-controlled study.

BACKGROUND & AIMS: To investigate the effects of probiotic in alleviation of stress in stressed adults, along our focus to identify and justify strain specificity on selected health benefits with a precisely targeted population. METHODS: This 12-weeks randomized, double-blind and placebo-controlled study investigated the effects of a probiotic (Lactobacillus plantarum P8; 10 log CFU daily) on psychological, memory and cognition parameters in one hundred and three (P8 n = 52, placebo n = 51) stressed adults with mean age of 31.7 ± 11.1 years old. All subjects fulfilled the criteria of moderate stress upon diagnosis using the PSS-10 questionnaire. RESULTS: At the end of study, subjects on P8 showed reduced scores of stress (mean difference 2.94; 95% CI 0.08 to 5.73; P = 0.048), anxiety (mean difference 2.82; 95% CI 0.35 to 5.30; P = 0.031) and total score (mean difference 8.04; 95% CI 0.73 to 15.30; P = 0.041) as compared to placebo after 4-weeks, as assessed by the DASS-42 questionnaire. Although plasma cortisol levels were only marginally different between placebo and P8 (mean difference 3.28 ug/dl; 95% CI -7.09 to 0.52; P = 0.090), pro-inflammatory cytokines such as IFN-γ (mean difference 8.07 pg/ml; 95% CI -11.2 to -4.93; P < 0.001) and TNF-α (mean difference 1.52 pg/ml; 95% CI -2.14 to -0.89; P < 0.001) showed higher reduction as compared to placebo over 12-weeks. These were accompanied by enhanced memory and cognitive traits such as social emotional cognition and verbal learning and memory upon administration of P8 as compared to the placebo, with different effects in women as compared to men. CONCLUSIONS: The present data illustrated that L. plantarum P8 is a feasible and natural intervention for the alleviation of selected stress, anxiety, memory and cognitive symptoms in stressed adults. TRIAL REGISTRATION: Approved by the JEPeM-USM Review Panel on Clinical Studies (Approval number USM/JEPeM/16050195) and was registered at ClinicalTrials.gov (identifier number NCT03268447).

Lactobacillus Strains Alleviated Aging Symptoms and Aging-Induced Metabolic Disorders in Aged Rats.

Aging is an inevitable and ubiquitous progress that affects all living organisms. A total of 18 strains of lactic acid bacteria (LAB) were evaluated on the activation of adenosine monophosphate-activated protein kinase (AMPK), an intracellular energy sensor mediating lifespan extension. The cell-free supernatant (CFS) of Lactobacillus fermentum DR9 (LF-DR9), Lactobacillus paracasei OFS 0291 (LP-0291), and Lactobacillus helveticus OFS 1515 (LH-1515) showed the highest activation of AMPK and was further evaluated. The phosphorylation of AMPK by these three LAB strains was more evident in U2OS and C2C12 cells, compared to the other cell lines and control (P < .05). Using premature senescent Sprague-Dawley rats induced by D-galactose (D-gal), the administration of LAB (10 log CFU/rat/day) for 12 weeks prevented the shortening of telomere length in D-gal-treated rats compared to the untreated control (P < .05). LF-DR9 lowered gene expression of p53, a known senescent biomarker, in gastrocnemius muscle and tibia compared to the control. The selected LAB strains also enhanced lipid, renal, and liver profile of rats, suggesting added potential of the strains in preventing aging-induced metabolic diseases. Strain LP-0291 and LH-1515 showed ability to adhere to mucin, no antibiotic resistance, tolerated and proliferated under gastric and intestinal simulated conditions, and inhibited the growth of pathogens Escherichia coli, Staphylococcus aureus, and Staphylococcus epidermidis, comparable to commercial probiotic LF-DR9 and Lactobacillus sakei Probio 65. This study provided an insight into the potential of LAB for exhibiting antisenescence effects, with potentials as new medicinal foods for targeted antiaging therapies.

Lactobacillus plantarum DR7 Reduces Cholesterol via Phosphorylation of AMPK That Down-regulated the mRNA Expression of HMG-CoA Reductase.

Hypercholesterolemia is one of the primary risk factors for cardiovascular diseases. The use of lactobacilli probiotics to reduce blood cholesterol levels have been extensively reported. However, more information is needed to evaluate the possible mechanisms involved and to identify possible targets for further therapeutic development. In this study, strains of lactobacilli were screened based on the ability to assimilate cholesterol, and prevention of cholesterol accumulation in hepatic (HepG2) and intestinal (HT-29) cells. Cell free supernatant (CFS) from Lactobacillus plantarum DR7 showed a higher ability to assimilate cholesterol, reduction in cholesterol accumulation in both HepG2 and HT-29 cells, accompanied by reduced mRNA expression of HMG-CoA reductase (HMGCR) in HepG2 (p<0.05), compared to other lactobacilli. The reduction of HMGCR expression was also diminished in the presence of an AMPK inhibitor (Compound C), suggesting that L. plantarum DR7 exerted its effect via the AMPK pathway, typically via the phosphorylation of AMPK instead of the AMPK mRNA expression in HepG2 (p<0.05). Altogether, our present study illustrated that lactobacilli could exert cholesterol lowering properties along the AMPK pathway, specifically via phosphorylation of AMPK that led to reduced expression of HMGCR.

Development and validation of a Chinese translated questionnaire: A single simultaneous tool for assessing gastrointestinal and upper respiratory tract related illnesses in pre-school children.

OBJECTIVES: Children are prone to contagious illnesses that come from peers in nurseries, kindergartens, and day care centres. The administration of probiotics has been reported to decrease the episodes of such illnesses, leading to decreased absences and consumption of antibiotics. With less emphasis on, and preferences for, blood collection from young subjects, quantifiable data are merely obtained from surveys and questionnaires. Malaysia has a population which is 25% ethnic Chinese. We aimed to develop a single tool that enables simultaneous assessments of both gastrointestinal and respiratory tract-related illnesses among young Chinese children. METHODS: The English-language validated questionnaires using data about demographics and monthly health records were translated into the Chinese language. Both forward and backward translated versions were validated. RESULTS: The developed demographic and monthly health questionnaires showed an overall item-level content validity index (I-CVI) of 0.99 and 0.97, respectively; while the translated Chinese versions showed I-CVI of 0.97 and 0.98, respectively. Item-level of response process validity index of 1.00 for this questionnaire was obtained from 30 respondents inferring that the items were clear and comprehensible. CONCLUSIONS: This study showed acceptable levels validity in the Chinese translated version, illustrating a valid and reliable tool to be used for simultaneous assessment of gastrointestinal and respiratory tract-related illnesses in young children that is applicable for Malaysia's Chinese population and other Chinese-speaking nations.