Alkaline Phosphatase-Mimicking Peptide Nanofibers for Osteogenic Differentiation.

Recognition of molecules and regulation of extracellular matrix synthesis are some of the functions of enzymes in addition to their catalytic activity. While a diverse array of enzyme-like materials have been developed, these efforts have largely been confined to the imitation of the chemical structure and catalytic activity of the enzymes, and it is unclear whether enzyme-mimetic molecules can also be used to replicate the matrix-regulatory roles ordinarily performed by natural enzymes. Self-assembled peptide nanofibers can provide multifunctional enzyme-mimetic properties, as the active sequences of the target enzymes can be directly incorporated into the peptides. Here, we report enhanced bone regeneration efficiency through peptide nanofibers carrying both catalytic and matrix-regulatory functions of alkaline phosphatase, a versatile enzyme that plays a critical role in bone formation by regulating phosphate homeostasis and calcifiable bone matrix formation. Histidine presenting peptide nanostructures were developed to function as phosphatases. These molecules are able to catalyze phosphate hydrolysis and serve as bone-like nodule inducing scaffolds. Alkaline phosphatase-like peptide nanofibers enabled osteogenesis for both osteoblast-like and mesenchymal cell lines.

Effect of human umbilical cord stem cells (HUMSC) administration on collagen expression in the anterior vaginal wall in menopausal rats.

Objective: To evaluate the outcome of human umbilical cord stem cells (HUMSC) administration on collagen expression within the frontal vaginal wall of menopausal rats. Materials and Methods: We conducted an experimental, randomized post-test-only controlled group design. The study samples were 40 healthy female Winstar rat with the age of 8-12 weeks that had been ovariectomized, had never mated, and weighed 18-22 grams. The umbilical cord was obtained from voluntary donors who did not have a history of hepatitis B, hepatitis C, HIV, cytomegalovirus infection, treponema pallidum infection, or a history of other infections transmitted through the blood, placental tract, and genitals. Data collection (frontal vaginal wall of the rat) was carried out in a controlled environment with the consideration that all conditions were maintained equally and could be controlled. Results: There were 36 samples. A total of 13 menopausal rats (72%) had strong collagen expression and 5 rats had weak-to-moderate collagen expression (28%). On the other hand, 18 menopausal rats (100%) that belonged to the control group had weak-moderate collagen expression, and no menopausal rats appeared to have strong expression (0%). The administration of collagen to the anterior vaginal wall of postmenopausal rats proved to be effective by increasing the strong collagen expression in the damaged anterior vagina of postmenopausal female rats (p<0.05). Conclusion: Administration of HUMSC resulted in an increase in collagen levels in the anterior vaginal tissue of postmenopausal female rats. These results demonstrate significant therapeutic potential for the treatment of pelvic floor dysfunction.

Antibiotics susceptibility of Escherichia coli isolates from clinical specimens before and during COVID-19 pandemic.

BACKGROUND AND OBJECTIVES: Escherichia coli is a Gram-negative organism causing mild to severe infections, with a wide spectrum range of organs involved. The study aimed to describe antibiotics susceptibility of E. coli from clinical specimens from October 11, 2019 to September 11, 2020. MATERIALS AND METHODS: Study was conducted retrospectively in a private microbiology laboratory in Mataram Indonesia. Period of study divided as two groups after WHO declared COVID-19 as pandemic by March 11, 2020; group A including the specimen related to September 2019 to March 11th 2020 and group B including the specimens related to March 11th 2020 to September 2020. All clinical specimens were subjected to identify E. coli isolates and their antibiotics susceptibility using WHO-NET 5.6 version. RESULTS: Totally, 148 E. coli isolates were found in group A and 62 isolates in group B. Prevalence of extended-spectrum beta lactamase (ESBL)- producing E. coli in group A was 50% and in group B was 20.9% with significantly difference (p<0.05). There was an increase in susceptibility to 10/16 antibiotics; where 3 antibiotics ofloxacin, aztreonam, and fosfomycin were significant (p<0.05). There was a significant decrease in susceptibility to the antibiotics piperacillin (p=0.012), amoxicillin (p=0.002), cefadroxil (p=0.036) and ampicillin (p=0.036). Type of infections between two groups: musculoskeletal infections, pneumonia, urinary tract infections and sepsis were not significant. CONCLUSION: Reduced number of E. coli isolates between two groups with decrease of ESBL-producing E. coli contribute in dynamics of antibiotics susceptibility. The longer period of analysis is needed to be done, due to the ongoing COVID-19 pandemic.

Association of Apolipoprotein E Gene Polymorphism with Lipid Profile and Ischemic Stroke Risk in Type 2 Diabetes Mellitus Patients.

BACKGROUND: Altered lipid profiles have consistently been linked to cerebrovascular events. Ischemic stroke (IS) was a common comorbid condition established in type 2 diabetes mellitus (T2DM). The apolipoprotein E (ApoE) gene which has a notably critical function in lipoprotein metabolism is believed as one of the potential candidate genes susceptible to IS complications in T2DM. This research aimed to determine the association of apolipoprotein E gene polymorphism with lipid profile and IS risk in T2DM patients. METHODS: This case-control study involved a total of 60 diabetic participants divided into two groups with and without IS. ApoE was genotyped using PCR and sequencing analysis. RESULTS: The most predominant genotype observed in 27 participants (45%) was E3/E3. Lower levels of high-density lipoprotein cholesterol (HDL-C) were found in ε2 carriers (p=0.003; 95% CI -23.35--4.89) and ε4 carriers (p=0.019; 95% CI 1.38-14.55) compared to ε3 homozygotes. Total cholesterol (TC), triglyceride, and low-density lipoprotein cholesterol (LDL-C) levels had no association with ApoE gene polymorphism in this study. ApoE gene polymorphism was not related to IS in T2DM (p=0.06; adjusted OR: 4.71; 95% CI 0.93-23.79). CONCLUSIONS: ApoE ε2 and ε4 carriers were associated with lower levels of HDL-C. No association was identified between ApoE gene polymorphism and IS in T2DM patients.

Effect of dietary protein regime on meat quality traits and carcase nutrient content of broilers from two commercial genotypes.

UNLABELLED: 1. The present study aimed to evaluate breast muscle quality and muscle myofibre density in two broiler genotypes given three dietary protein feeding regimes. 2. A total of 1440 Ross 308 and 708 broilers (720/genotype) were given one of three dietary protein regimes (Low: 192, 166 and 155 g/kg, CONTROL: 229, 199 and 182 g/kg and High: 266, 235 and 207 g/kg for starter, grower and finisher diets respectively). On d 21 and 42, initial protein solubility, DNA and myofibre density were determined. On d 42, breast meat samples were collected for meat quality analyses. 3. There were no genotype effects on pH(24), L*, a* or b* values. Thawing loss was higher in meat from the 708 than the 308 genotype. 4. Broilers given the low protein feeding regime had highest carcase fat, lowest pH(24) and palest meat. The low regression coefficients between dietary protein content and changes in pH(24) and lightness, however, demonstrated that dietary protein was not the principal factor influencing these traits. 5. Although sarcoplasmic protein solubility was similar among protein feeding regimes on d 21, the highest sarcoplasmic protein solubility on d 42 was in broilers on the high protein regime. 6. Myofibre density was highest in meat from broilers given the low protein regime. 7. It was concluded that dietary-protein content contributes significantly to both protein solubility and myofibre density.

Sequential secretion of collagenolytic, elastolytic, and keratinolytic proteases in peptide-limited cultures of two Bacillus cereus strains isolated from wool.

AIMS: To characterize the secretion of proteolytic activities against keratin, collagen and elastin in liquid cultures of Bacillus cereus IZ-06b and IZ-06r isolated from wool. METHODS AND RESULTS: Growth of B. cereus IZ-06b and IZ-06r were characterized in batch culture. Both strains needed an organic nitrogen source, were able to grow on wool or peptone as sole carbon and nitrogen sources, and metabolized glucose, maltose and other simple sugars. Proteolytic activities were investigated in batch cultures grown in peptide-restricted, carbon-sufficient medium. Secretion of proteases was induced by peptide limitation while different proteolytic activities appeared sequentially in the growth medium. When the most available components of the peptone were depleted, collagenolytic and elastolytic proteases were produced. These were later replaced by the production of keratinolytic protease. CONCLUSIONS: B. cereus can adjust its proteolytic affinity profile in response to the supply of organic nitrogen and sequentially secrete proteases with activities targeted against increasingly inaccessible proteinous substrates as the nutritional availability in the environment deteriorates. SIGNIFICANCE AND IMPACT OF THE STUDY: Peptide-limited, carbon-sufficient growth media containing no proteinous substrates are well suited for protease production in B. cereus while growth conditions can be adjusted to optimize the proteolytic affinity profiles.

Light-Triggered Drug Release from 3D-Printed Magnetic Chitosan Microswimmers.

Advances in design and fabrication of functional micro/nanomaterials have sparked growing interest in creating new mobile microswimmers for various healthcare applications, including local drug and other cargo ( e. g., gene, stem cell, and imaging agent) delivery. Such microswimmer-based cargo delivery is typically passive by diffusion of the cargo material from the swimmer body; however, controlled active release of the cargo material is essential for on-demand, precise, and effective delivery. Here, we propose a magnetically powered, double-helical microswimmer of 6 µm diameter and 20 µm length that can on-demand actively release a chemotherapeutic drug, doxorubicin, using an external light stimulus. We fabricate the microswimmers by two-photon-based 3D printing of a natural polymer derivative of chitosan in the form of a magnetic polymer nanocomposite. Amino groups presented on the microswimmers are modified with doxorubicin by means of a photocleavable linker. Chitosan imparts the microswimmers with biocompatibility and biodegradability for use in a biological setting. Controlled steerability of the microswimmers is shown under a 10 mT rotating magnetic field. With light induction at 365 nm wavelength and 3.4 × 10-1 W/cm2 intensity, 60% of doxorubicin is released from the microswimmers within 5 min. Drug release is ceased by controlled patterns of light induction, so as to adjust the desired release doses in the temporal domain. Under physiologically relevant conditions, substantial degradation of the microswimmers is shown in 204 h to nontoxic degradation products. This study presents the combination of light-triggered drug delivery with magnetically powered microswimmer mobility. This approach could be extended to similar systems where multiple control schemes are needed for on-demand medical tasks with high precision and efficiency.

Angiogenic peptide nanofibers repair cardiac tissue defect after myocardial infarction.

Myocardial infarction remains one of the top leading causes of death in the world and the damage sustained in the heart eventually develops into heart failure. Limited conventional treatment options due to the inability of the myocardium to regenerate after injury and shortage of organ donors require the development of alternative therapies to repair the damaged myocardium. Current efforts in repairing damage after myocardial infarction concentrates on using biologically derived molecules such as growth factors or stem cells, which carry risks of serious side effects including the formation of teratomas. Here, we demonstrate that synthetic glycosaminoglycan (GAG) mimetic peptide nanofiber scaffolds induce neovascularization in cardiovascular tissue after myocardial infarction, without the addition of any biologically derived factors or stem cells. When the GAG mimetic nanofiber gels were injected in the infarct site of rodent myocardial infarct model, increased VEGF-A expression and recruitment of vascular cells was observed. This was accompanied with significant degree of neovascularization and better cardiac performance when compared to the control saline group. The results demonstrate the potential of future clinical applications of these bioactive peptide nanofibers as a promising strategy for cardiovascular repair. STATEMENT OF SIGNIFICANCE: We present a synthetic bioactive peptide nanofiber system can enhance cardiac function and enhance cardiovascular regeneration after myocardial infarction (MI) without the addition of growth factors, stem cells or other biologically derived molecules. Current state of the art in cardiac repair after MI utilize at least one of the above mentioned biologically derived molecules, thus our approach is ground-breaking for cardiovascular therapy after MI. In this work, we showed that synthetic glycosaminoglycan (GAG) mimetic peptide nanofiber scaffolds induce neovascularization and cardiomyocyte differentiation for the regeneration of cardiovascular tissue after myocardial infarction in a rat infarct model. When the peptide nanofiber gels were injected in infarct site at rodent myocardial infarct model, recruitment of vascular cells was observed, neovascularization was significantly induced and cardiac performance was improved. These results demonstrate the potential of future clinical applications of these bioactive peptide nanofibers as a promising strategy for cardiovascular repair.

Analysis of specific antibody and cellular immune response to first-dose measles vaccine Edmonston-Zagreb in 9-month-old infants

Background: Measles vaccinations have been suggested to provide immune protection and decreased measles incidence. However, there was a limited study evaluating how the measles vaccine elicits specific immune responses. Objective: This study aimed to evaluate both humoral and cellular immunity to first-dose measles vaccine Edmonston-Zagreb (EZ) in 9-month-old Indonesian infants. Methods: A cohort study was conducted on 9-month-old infants who got the first-dose of measles vaccine EZ. Measles-specific immunoglobulin G (IgG) antibody serum levels were measured using plaque-reduction microneutralization assay. Peripheral blood mononuclear cells were stimulated with a measles-specific peptide to identify a cellular immune response. Quantification of CD4+ and CD8+ T-cells producing interferon-gamma (IFN-ɣ) and interleukin 17-A (IL-17A) were conducted by flow cytometry. Humoral and cellular immune response parameters were analyzed over time. Results: The prevalence of seropositivity rates was 85.8% at 1-month after vaccination and 16.67% at 6-months postvaccination. Measles-specific IgG antibodies increased significantly at 1-month after measles vaccination. However, they decreased significantly 6-months after vaccination. IFN-ɣ and IL-17A secreting T-cells increased significantly at 1-month after measles vaccination. Interestingly, a significant decrease of IFN-ɣ and IL-17A secreting CD4+ T cells was noticed 6-months postvaccination compared to IFN-ɣ and IL-17A secreting CD8+ T cells. Conclusion: Our study suggests that the first-dose measles vaccine on 9-months-old infants seems to induce both humoral and cellular immune responses that decline 6-months after vaccination (AU)

Characterization and antimicrobial performance of nano silver coatings on leather materials.

In this study, the characterization and the antimicrobial properties of nano silver (nAg) coating on leather were investigated. For this purpose, turbidity, viscosity and pH of nAg solutions prepared by the sol-gel method were measured. The formation of films from these solutions was characterized according to temperature by Differential Thermal Analysis-Thermogravimetry (DTA-TG) equipment. The surface morphology of treated leathers was observed using Scanning Electron Microscopy (SEM). The antimicrobial performance of nAg coatings on leather materials to the test microorganisms as Escherichia coli , Staphylococcus aureus , Candida albicans and Aspergillius niger was evaluated by the application of qualitative (Agar overlay method) and quantitative (percentage of microbial reduction) tests. According to qualitative test results it was found that 20 µg/cm (2) and higher concentrations of nAg on the leather samples were effective against all microorganisms tested. Moreover, quantitative test results showed that leather samples treated with 20 µg/cm (2) of nAg demonstrated the highest antibacterial activity against E. coli with 99.25% bacterium removal, whereas a 10 µg/cm (2) concentration of nAg on leather was enough to exhibit the excellent percentage reduction against S. aureus of 99.91%. The results are promising for the use of colloidal nano silver solution on lining leather as antimicrobial coating.

Basal Lamina Mimetic Nanofibrous Peptide Networks for Skeletal Myogenesis.

Extracellular matrix (ECM) is crucial for the coordination and regulation of cell adhesion, recruitment, differentiation and death. Therefore, equilibrium between cell-cell and cell-matrix interactions and matrix-associated signals are important for the normal functioning of cells, as well as for regeneration. In this work, we describe importance of adhesive signals for myoblast cells' growth and differentiation by generating a novel ECM mimetic peptide nanofiber scaffold system. We show that not only structure but also composition of bioactive signals are important for cell adhesion, growth and differentiation by mimicking the compositional and structural properties of native skeletal muscle basal lamina. We conjugated laminin-derived integrin binding peptide sequence, "IKVAV", and fibronectin-derived well known adhesive sequence, "RGD", into peptide nanostructures to provide adhesive and myogenic cues on a nanofibrous morphology. The myogenic and adhesive signals exhibited a synergistic effect on model myoblasts, C2C12 cells. Our results showed that self-assembled peptide nanofibers presenting laminin derived epitopes support adhesion, growth and proliferation of the cells and significantly promote the expression of skeletal muscle-specific marker genes. The functional peptide nanofibers used in this study present a biocompatible and biodegradable microenvironment, which is capable of supporting the growth and differentiation of C2C12 myoblasts into myotubes.

Synthesis and antifungal activity of some new arylidenamino compounds.

A series of 3-alkyl/aryl-4-arylidenamimo-4,5-dihydro-1H-1,2,4- triazol-5-ones was synthesized and characterized by elemental and spectral analysis. These compound were tested for in vitro antibacterial and antifungal activity in solid agar cultures against eight microorganisms. In these test, some of the new compounds were shown to be very potent in vitro antifungal activity against the used fungi.

Antigenic variation of clade 2.1 H5N1 virus is determined by a few amino acid substitutions immediately adjacent to the receptor binding site.

UNLABELLED: Highly pathogenic avian influenza (HPAI) viruses of the H5N1 subtype are genetically highly variable and have diversified into multiple phylogenetic clades over the past decade. Antigenic drift is a well-studied phenomenon for seasonal human influenza viruses, but much less is known about the antigenic evolution of HPAI H5N1 viruses that circulate in poultry. In this study, we focused on HPAI H5N1 viruses that are enzootic to Indonesia. We selected representative viruses from genetically distinct lineages that are currently circulating and determined their antigenic properties by hemagglutination inhibition assays. At least six antigenic variants have circulated between 2003, when H5N1 clade 2.1 viruses were first detected in Indonesia, and 2011. During this period, multiple antigenic variants cocirculated in the same geographic regions. Mutant viruses were constructed by site-directed mutagenesis to represent each of the circulating antigenic variants, revealing that antigenic differences between clade 2.1 viruses were due to only one or very few amino acid substitutions immediately adjacent to the receptor binding site. Antigenic variants of H5N1 virus evaded recognition by both ferret and chicken antibodies. The molecular basis for antigenic change in clade 2.1 viruses closely resembled that of seasonal human influenza viruses, indicating that the hemagglutinin of influenza viruses from different hosts and subtypes may be similarly restricted to evade antibody recognition. IMPORTANCE: Highly pathogenic avian influenza (HPAI) H5N1 viruses are responsible for severe outbreaks in both commercial and backyard poultry, causing considerable economic losses and regular zoonotic transmissions to humans. Vaccination is used increasingly to reduce the burden of HPAI H5N1 virus in poultry. Influenza viruses can escape from recognition by antibodies induced upon vaccination or infection through genetic changes in the hemagglutinin protein. The evolutionary patterns and molecular basis of antigenic change in HPAI H5N1 viruses are poorly understood, hampering formulation of optimal vaccination strategies. We have shown here that HPAI H5N1 viruses in Indonesia diversified into multiple antigenic variants, that antigenic differences were due to one or a very few substitutions near the receptor binding site, and that the molecular basis for antigenic change was remarkably similar to that for seasonal human influenza viruses. These findings have consequences for future vaccination and surveillance considerations and contribute to the understanding of the antigenic evolution of influenza viruses.

Characterization and antimicrobial performance of nano silver coatings on leather materials

In this study, the characterization and the antimicrobial properties of nano silver (nAg) coating on leather were investigated. For this purpose, turbidity, viscosity and pH of nAg solutions prepared by the sol-gel method were measured. The formation of films from these solutions was characterized according to temperature by Differential Thermal Analysis-Thermogravimetry (DTA-TG) equipment. The surface morphology of treated leathers was observed using Scanning Electron Microscopy (SEM). The antimicrobial performance of nAg coatings on leather materials to the test microorganisms as Escherichia coli, Staphylococcus aureus, Candida albicans and Aspergillius niger was evaluated by the application of qualitative (Agar overlay method) and quantitative (percentage of microbial reduction) tests. According to qualitative test results it was found that 20 μg/cm2 and higher concentrations of nAg on the leather samples were effective against all microorganisms tested. Moreover, quantitative test results showed that leather samples treated with 20 μg/cm2 of nAg demonstrated the highest antibacterial activity against E. coli with 99.25% bacterium removal, whereas a 10 μg/cm2 concentration of nAg on leather was enough to exhibit the excellent percentage reduction against S. aureus of 99.91%. The results are promising for the use of colloidal nano silver solution on lining leather as antimicrobial coating.

Identification of extracellular enzyme producing alkalophilic bacilli from Izmir province by 16S-ITS rDNA RFLP.

AIMS: To screen industrially important extracellular enzymes from the newly isolated alkalophilic bacilli and to characterize them by phenotypic and 16S-internal transcribed spacer (ITS) rDNA restriction pattern analysis. METHODS AND RESULTS: Three different environmental samples, soil, leather and horse faeces, were collected within the province of Izmir. Isolates grown on Horikoshi-I medium for 24 h at 37 degrees C were screened for extracellular enzyme activity by using eight different substrates: birchwood xylan, carboxymethylcellulose, casein, citrus pectin, polygalacturonic acid, soluble starch, and Tween 20 and 80. In total, 115 extracellular enzyme-producing bacilli were obtained. Casein was hydrolysed by 78%, soluble starch by 67%, citrus pectin by 63%, polygalacturonic acid by 62%, Tween 20 by 34%, birchwood xylan by 16%, Tween 80 by 12%, and carboxymethylcellulose by 3% of the isolates. The isolates were differentiated into 19 distinct homology groups by the 16S-ITS rDNA restriction pattern analysis. CONCLUSIONS: Eight different extracellular enzyme activities were determined in 115 endospore forming bacilli. The largest 16S-ITS rDNA homology group (HT1) included 36% of the isolates, 98% of which degraded casein, polygalacturonic acid, pectin and starch. SIGNIFICANCE AND IMPACT OF THE STUDY: This study is the first report on the characterization of the industrial enzyme-producing alkalophilic bacilli by 16S-ITS rDNA restriction fragment length polymorphism (RFLP). Restriction profiles of 64% of the isolates were found to be different from those of five reference strains used.