Phenotypic Frailty Assessment in Mice: Development, Discoveries, and Experimental Considerations.

The underlying mechanisms contributing to the onset of frailty, its progression, and its mortality risk remain unknown. Recently, the two most common human frailty assessments were reverse-translated to mice. Here, we highlight the development of the mouse frailty phenotype, unique discoveries, experimental considerations, and future perspectives.

Adenosine kinase attenuates cardiomyocyte microtubule stabilization and protects against pressure overload-induced hypertrophy and LV dysfunction.

Adenosine exerts numerous protective actions in the heart, including attenuation of cardiac hypertrophy. Adenosine kinase (ADK) converts adenosine to adenosine monophosphate (AMP) and is the major route of myocardial adenosine metabolism, however, the impact of ADK activity on cardiac structure and function is unknown. To examine the role of ADK in cardiac homeostasis and adaptation to stress, conditional cardiomyocyte specific ADK knockout mice (cADK-/-) were produced using the MerCreMer-lox-P system. Within 4 weeks of ADK disruption, cADK-/- mice developed spontaneous hypertrophy and increased ß-Myosin Heavy Chain expression without observable LV dysfunction. In response to 6 weeks moderate left ventricular pressure overload (transverse aortic constriction;TAC), wild type mice (WT) exhibited ~60% increase in ventricular ADK expression and developed LV hypertrophy with preserved LV function. In contrast, cADK-/- mice exhibited significantly greater LV hypertrophy and cardiac stress marker expression (atrial natrurietic peptide and ß-Myosin Heavy Chain), LV dilation, reduced LV ejection fraction and increased pulmonary congestion. ADK disruption did not decrease protein methylation, inhibit AMPK, or worsen fibrosis, but was associated with persistently elevated mTORC1 and p44/42 ERK MAP kinase signaling and a striking increase in microtubule (MT) stabilization/detyrosination. In neonatal cardiomyocytes exposed to hypertrophic stress, 2-chloroadenosine (CADO) or adenosine treatment suppressed MT detyrosination, which was reversed by ADK inhibition with iodotubercidin or ABT-702. Conversely, adenoviral over-expression of ADK augmented CADO destabilization of MTs and potentiated CADO attenuation of cardiomyocyte hypertrophy. Together, these findings indicate a novel adenosine receptor-independent role for ADK-mediated adenosine metabolism in cardiomyocyte microtubule dynamics and protection against maladaptive hypertrophy.

Role of bone marrow-derived CD11c+ dendritic cells in systolic overload-induced left ventricular inflammation, fibrosis and hypertrophy.

Inflammatory responses play an important role in the development of left ventricular (LV) hypertrophy and dysfunction. Recent studies demonstrated that increased T-cell infiltration and T-cell activation contribute to LV hypertrophy and dysfunction. Dendritic cells (DCs) are professional antigen-presenting cells that orchestrate immune responses, especially by modulating T-cell function. In this study, we investigated the role of bone marrow-derived CD11c+ DCs in transverse aortic constriction (TAC)-induced LV fibrosis and hypertrophy in mice. We observed that TAC increased the number of CD11c+ cells and the percentage of CD11c+ MHCII+ (major histocompatibility complex class II molecule positive) DCs in the LV, spleen and peripheral blood in mice. Using bone marrow chimeras and an inducible CD11c+ DC ablation model, we found that depletion of bone marrow-derived CD11c+ DCs significantly attenuated LV fibrosis and hypertrophy in mice exposed to 24 weeks of moderate TAC. CD11c+ DC ablation significantly reduced TAC-induced myocardial inflammation as indicated by reduced myocardial CD45+ cells, CD11b+ cells, CD8+ T cells and activated effector CD8+CD44+ T cells in LV tissues. Moreover, pulsing of autologous DCs with LV homogenates from TAC mice promoted T-cell proliferation. These data indicate that bone marrow-derived CD11c+ DCs play a maladaptive role in hemodynamic overload-induced cardiac inflammation, hypertrophy and fibrosis through the presentation of cardiac self-antigens to T cells.

Cardiomyocyte dimethylarginine dimethylaminohydrolase-1 (DDAH1) plays an important role in attenuating ventricular hypertrophy and dysfunction.

Asymmetric dimethylarginine (ADMA) is an endogenous inhibitor of nitric oxide synthases that limits nitric oxide bioavailability. Dimethylarginine dimethylaminohydrolase-1 (DDAH1) exerts a critical role for ADMA degradation and plays an important role in NO signaling. In the heart, DDAH1 is observed in endothelial cells and in the sarcolemma of cardiomyocytes. While NO signaling is important for cardiac adaptation to stress, DDAH1 impact on cardiomyocyte homeostasis is not clear. Here we used the MerCreMer-LoxP model to specifically disrupt cardiomyocyte DDAH1 expression in adult mice to determine the physiological impact of cardiomyocyte DDAH1 under basal conditions and during hypertrophic stress imposed by transverse aortic constriction (TAC). Under control conditions, cardiomyocyte-specific DDAH1 knockout (cDDAH KO) had no detectable effect on plasma ADMA and left ventricular (LV) hypertrophy or function in adult or aging mice. In response to TAC, DDAH1 levels were elevated 2.5-fold in WT mice, which exhibited no change in LV or plasma ADMA content and moderate LV hypertrophy and LV dysfunction. In contrast, cDDAH1 KO mice exposed to TAC showed no increase in LV DDAH1 expression, slightly increased LV tissue ADMA levels, no increase in plasma ADMA, but significantly exacerbated LV hypertrophy, fibrosis, nitrotyrosine production, and LV dysfunction. These findings indicate cardiomyocyte DDAH1 activity is dispensable for cardiac function under basal conditions, but plays an important role in attenuating cardiac hypertrophy and ventricular remodeling under stress conditions, possibly through locally confined regulation of subcellular ADMA and NO signaling.

Double-stranded RNA-dependent protein kinase deficiency protects the heart from systolic overload-induced congestive heart failure.

BACKGROUND: Double-stranded RNA-dependent protein kinase (PKR) is a eukaryotic initiation factor 2α kinase that inhibits mRNA translation under stress conditions. PKR also mediates inflammatory and apoptotic signaling independently of translational regulation. Congestive heart failure is associated with cardiomyocyte hypertrophy, inflammation, and apoptosis, but the role of PKR in left ventricular hypertrophy and the development of congestive heart failure has not been examined. METHODS AND RESULTS: We observed increased myocardial PKR expression and translocation of PKR into the nucleus in humans and mice with congestive heart failure. To determine the impact of PKR on the development of congestive heart failure, PKR knockout and wild-type mice were exposed to pressure overload produced by transverse aortic constriction. Although heart size increased similarly in wild-type and PKR knockout mice after transverse aortic constriction, PKR knockout mice exhibited very little pulmonary congestion, well-preserved left ventricular ejection fraction and contractility, and significantly less myocardial fibrosis compared with wild-type mice. Bone marrow-derived cells from wild-type mice did not abolish the cardiac protective effect observed in PKR knockout mice, whereas bone marrow-derived cells from PKR knockout mice had no cardiac protective effect in wild-type mice. Mechanistically, PKR knockout attenuated transverse aortic constriction-induced tumor necrosis factor-α expression and leukocyte infiltration and lowered cardiac expression of proapoptotic factors (Bax and caspase-3), so that PKR knockout hearts were more resistant to transverse aortic constriction-induced cardiomyocyte apoptosis. PKR depletion in isolated cardiomyocytes also conferred protection against tumor necrosis factor-α- or lipopolysaccharide-induced apoptosis. CONCLUSION: PKR is a maladaptive factor upregulated in hemodynamic overload that contributes to myocardial inflammation, cardiomyocyte apoptosis, and the development of congestive heart failure.

Electroreductive Access to 1,2-Aminoalcohols via Cross Aza-Pinacol Coupling of N-Acyl Diarylketimines and Aldehydes.

We present highly efficient and operationally simple synthetic methods for 1,2-aminoalcohols via electroreductive cross aza-pinacol coupling between N-acyl diarylketimines and aldehydes. Preliminary mechanistic studies including cyclic voltammetry and density functional theory (DFT) calculations suggest that the reaction is instigated by selective electrochemical single electron transfer (SET) of N-acylketimines. The developed electrochemical protocol is compatible to biorelevant functional groups, enabling late-stage functionalization of pharmacophores.

pH-sustaining nanostructured hydroxyapatite/alginate composite hydrogel for gastric protection and intestinal release of Lactobacillus rhamnosusGG.

The gut microbiome is closely linked to gastrointestinal health and disease status. Oral administration of known probiotic strains is now considered a promising therapeutic strategy, especially for refractory diseases such as inflammatory bowel disease. In this study, we developed a nanostructured hydroxyapatite/alginate (HAp/Alg) composite hydrogel that protects its encapsulated probiotic Lactobacillus rhamnosus GG (LGG) by neutralizing hydrogen ions that penetrate the hydrogel in a stomach without inhibiting LGG release in an intestine. Surface and transection analyses of the hydrogel revealed characteristic patterns of crystallization and composite-layer formation. TEM revealed the dispersal of the nanosized HAp crystals and encapsulated LGG in the Alg hydrogel networks. The HAp/Alg composite hydrogel maintained its internal microenvironmental pH, thereby enabling the LGG to survive for substantially longer. At intestinal pH, the encapsulated LGG was completely released upon disintegration of the composite hydrogel. In a dextran sulfate sodium-induced colitis mouse model, we then assessed the therapeutic effect of the LGG-encapsulating hydrogel. This achieved intestinal delivery of LGG with minimal loss of enzymatic function and viability, ameliorating colitis by reducing epithelial damage, submucosal edema, inflammatory cell infiltration, and the number of goblet cells. These findings reveal the HAp/Alg composite hydrogel as a promising intestinal-delivery platform for live microorganisms including probiotics and live biotherapeutic products.

Ionically bridged dexamethasone sodium phosphate-zinc-PLGA nanocomplex in alginate microgel for the local treatment of ulcerative colitis.

Colon-targeted oral drug delivery systems comprising nanoparticles and microparticles have emerged as promising tools for the treatment of ulcerative colitis (UC) because they minimize side effects and maximize the local drug concentration. Dexamethasone sodium phosphate (DSP) is a potent anti-inflammatory glucocorticoid used for the treatment of UC. However, it remains a rather short-term treatment option owing to its side effects. In the present study, we developed the alginate gel encapsulating ionically bridged DSP-zinc-poly(lactic-co-glycolic acid) (PLGA) nanocomplex (DZP-NCs-in-microgel) for the oral local treatment of UC. The successful encapsulation of DSP-zinc-PLGA nanocomplex (DZP-NCs) in alginate microgel was confirmed by SEM imaging. The prepared gel released DZP-NCs in the stimulated intestinal fluid and dampened the release of DSP in the upper gastrointestinal tract. Furthermore, DZP-NCs-in-microgel alleviated colonic inflammation in a mouse model of dextran sodium sulfate-induced colitis by relieving clinical symptoms and histological marks. Our results suggest a novel approach for the oral colon-targeted delivery of dexamethasone sodium phosphate for the treatment of UC.

5-Fluorouracil crystal-incorporated, pH-responsive, and release-modulating PLGA/Eudragit FS hybrid microparticles for local colorectal cancer-targeted chemotherapy.

5-Fluorouracil (5-FU) is a widely used chemotherapeutic agent for colorectal cancer (CRC) owing to its potent anticancer effects. However, severe systemic side effects and poor drug accumulation in the CRC tissues limit its efficacy. This study aimed to develop 5-FU crystal-incorporated, pH-responsive, and release-modulating poly(d,l-lactide-co-glycolide)/Eudragit FS hybrid microparticles (5FU-EPMPs) for the local CRC-targeted chemotherapy. Approximately 150 µm 5FU-EPMPs were fabricated via the S/O/W emulsion solvent evaporation method, with 7.93 ± 0.24% and 87.23 ± 2.64% 5-FU loading and encapsulation efficiencies, respectively. Drug release profiles in a simulated pH environment of the gastrointestinal tract revealed that premature 5-FU release in the stomach and small intestine was prevented, thereby minimizing systemic 5-FU absorption. After reaching the colon, 5-FU was continuously released for >15 h, allowing long-term exposure of CRC tissues to sufficient 5-FU concentrations. Furthermore, in a CRC mouse model, the 5FU-EPMPs showed potent inhibition of tumor growth without signs of systemic toxicity. Thus, the 5FU-EPMPs represent a promising drug delivery system for local CRC-targeted chemotherapy.

On-demand reconstitutable hyaluronic acid-doped azathioprine microcrystals effectively ameliorate ulcerative colitis via selective accumulation in inflamed tissues.

Although CD44-targeted delivery of pure drug microcrystals of azathioprine (AZA) could be a desirable approach to treat ulcerative colitis (UC), premature drug release and systemic absorption before reaching the colitis region remain a major obstacle. In this study, to overcome these limitations, we developed on-demand reconstitutable HA-doped AZA microcrystals (EFS/HA-AZAs) via incorporating hyaluronic acid (HA)-doped AZA microcrystals (HA-AZAs) into a Eudragit FS (EFS) microcomposite. Since EFS acts as a protective layer, the premature release of AZA in the simulated conditions of the stomach and small intestine was substantially reduced, while HA-AZAs were successfully reconstituted from the EFS/HA-AZAs in the colonic environment, resulting from the pH-triggered dissolution of EFS. After complete reconstitution of HA-AZAs in the colon, HA-AZAs selectively accumulated in the inflamed region via the HA-CD44 interaction. Owing to successful colitis-targeted delivery, EFS/HA-AZAs showed potent anti-inflammatory effects in a dextran sulfate sodium-induced murine colitis model within 7 days without systemic toxicity. These results suggest that EFS/HA-AZAs could be a promising drug delivery system for UC treatment.

Cupriavidus necator-Produced Polyhydroxybutyrate/Eudragit FS Hybrid Nanoparticles Mitigates Ulcerative Colitis via Colon-Targeted Delivery of Cyclosporine A.

Polyhydroxybutyrate (PHB) has emerged as a novel material for replacing various plastics used in the medical field. However, its application as a drug-delivery carrier for colitis-targeted delivery has not been explored. In this study, we used biosynthesized PHB combined with Eudragit FS (EFS) and cyclosporine A (CSA) to develop pH-responsive controlled CSA-releasing nanoparticles (CSA-PENPs) for colitis-targeted drug delivery and demonstrated its enhanced therapeutic efficacy in a dextran sulfate sodium (DSS)-induced murine colitis model. PHB was successfully biosynthesized in the bacterium Cupriavidus necator, as demonstrated by 1H-NMR and FT-IR analyses. CSA-PENPs were fabricated via the oil-in-water emulsion solvent evaporation method. Owing to the potent pH-responsive and sustained drug release properties provided by PHB and EFS, CSA-PENPs could deliver a sufficient amount of CSA to inflamed tissues in the distal colon; in contrast, CSA-loaded EFS nanoparticles displayed premature burst release before reaching the target site. Due to enhanced CSA delivery to colitis tissues, CSA-PENPs exhibited potent anti-inflammatory effects in the DSS-induced murine colitis model. Overall, CSA-PENPs could be a promising drug-delivery system for treating ulcerative colitis.

Hyaluronic Acid-Conjugated PLGA Nanoparticles Alleviate Ulcerative Colitis via CD44-Mediated Dual Targeting to Inflamed Colitis Tissue and Macrophages.

Although various local anti-inflammatory therapies for ulcerative colitis have been developed, rapid drug elimination from inflamed colitis tissue and off-target side effects reduce their therapeutic efficacy. In this study, we synthesized curcumin (Cur)-loaded hyaluronic acid (HA)-conjugated nanoparticles (Cur-HA-PLGA-NPs) that target inflamed colitis tissue via HA-CD44 interaction with resident colonic epithelial cells and subsequently target activated macrophages for ulcerative colitis therapy. The synthesized spherical Cur-HA-PLGA-NPs showed physicochemical properties similar to those of non-HA-conjugated Cur-PLGA-NPs. HA-PLGA-NPs exhibited selective accumulation in inflamed colitis tissue with minimal accumulation in healthy colon tissue. HA functionalization enhanced targeted drug delivery to intestinal macrophages, significantly increasing HA-PLGA-NP cellular uptake. Importantly, the rectal administration of Cur-HA-PLGA-NPs exhibited better therapeutic efficacy than Cur-PLGA-NPs in animal studies. Histological examination revealed that Cur-HA-PLGA-NPs reduced inflammation with less inflammatory cell infiltration and accelerated recovery with re-epithelialization signs. Our results suggest that Cur-HA-PLGA-NPs are a promising delivery platform for treating ulcerative colitis.

Frailty: Past, present, and future?

The prevalence of frailty across the world in older adults is increasing dramatically and having frailty places a person at increased risk for many adverse health outcomes, including impaired mobility, falls, hospitalizations, and mortality. Globally, the concept of frailty is gaining attention and the scientific field has made great strides in identifying and conceptually defining frailty through consensus conferences, in advancing the overall science of frailty by drawing on basic science discoveries including concepts surrounding the hallmarks of aging, resilience, and intrinsic capacities, and in identifying the many challenges faced by professionals within diverse clinical settings. Currently, it is thought that frailty is preventable, thus the identification of a person's degree of frailty is vital. Identification of frailty is achievable through widely used frailty screening tools, which are valid, reliable, and easy to use. Following the identification of a person's degree of frailty, targeted intervention strategies, such as physical activity programs must be implemented. In this perspective, we provide a historical perspective of the frailty field since the last quarter of the 20th century to present. We identify the proposed underlying pathophysiology of multiple physiological systems, including compromised homeostasis and resilience. Next, we outline the available screening tools for frailty with a physical performance assessment and highlight specific benefits of physical activity. Lastly, we discuss current scientific evidence supporting the physical activity recommendations for the aging population and for older adults with frailty. The goal is to emphasize early detection of frailty and stress the value of physical activity.

Diethylenetriamine/NONOate-doped alginate hydrogel with sustained nitric oxide release and minimal toxicity to accelerate healing of MRSA-infected wounds.

This study demonstrates the development of a nitric oxide (NO)-releasing hydrogel wound dressing and its efficacy at accelerating methicillin-resistant Staphylococcus aureus (MRSA)-infected wound healing. A DETA/NONOate-doped alginate (Alg-DETA/NO) hydrogel was synthesized using alginate as a hydrogel-forming wound dressing material and diethylenetriamine/diazeniumdiolate (DETA/NONOate) as an NO donor. Alg-DETA/NO exhibited a prolonged NO release profile over a period of 4 days. The rheological properties of Alg-DETA/NO did not differ significantly from those of pure alginate. Importantly, Alg-DETA/NO showed potent antibacterial activity against MRSA, with minimal toxicity to mouse fibroblasts. The application of Alg-DETA/NO to MRSA-infected wounds in a mouse model showed a favorable wound healing with accelerated wound-size reduction and reduced skin bacterial infection. Additionally, histological examination revealed that Alg-DETA/NO reduced inflammation at the wound site and promoted re-epithelialization, angiogenesis, and collagen deposition. Thus, Alg-DETA/NO presented herein could serve as a safe and potent hydrogel dressing for the treatment of MRSA-infected wounds.

Triolein emulsion enhances temozolomide brain delivery: an experimental study in rats.

PURPOSE: To evaluate the enhancement of temozolomide (TMZ) delivery in the rat brain using a triolein emulsion. MATERIALS AND METHODS: Rats were divided into the five groups as following: group 1 (negative control), group 2 (treated with triolein emulsion and TMZ 20 mg/kg), and group 3 (TMZ 20 mg/kg treatment without triolein), group 4 (treated with triolein emulsion and TMZ 10 mg/kg), and group 5 (TMZ 10 mg/kg treatment without triolein). Triolein emulsion was infused into the right common carotid artery. One hour later, the TMZ concentration was evaluated quantitatively and qualitatively using high-performance liquid chromatography (HPLC-MS) and desorption electrospray ionization mass spectrometry (DESI-MS) imaging, respectively. The concentration ratios of the ipsilateral to contralateral hemisphere in each group were determined and the statistical analysis was conducted using an unpaired t-test. RESULTS: Quantitatively, the TMZ concentration ratio of the ipsilateral to the control hemisphere was 2.41 and 1.13 in groups 2 and 3, and were 2.49 and 1.14 in groups 4 and 5, respectively. Thus, the TMZ signal intensities of TMZ in group 2 and 4 were statistically high in the ipsilateral hemispheres. Qualitatively, the signal intensity of TMZ was remarkably high in the ipsilateral hemisphere in group 2 and 4. CONCLUSIONS: The triolein emulsion efficiently opened the blood-brain barrier and could provide a potential new strategy to enhance the therapeutic effect of TMZ. HPLC-MS and DESI-MS imaging were shown to be suitable for analyses of enhancement of brain TMZ concentrations.

Tumor-Penetrable Nitric Oxide-Releasing Nanoparticles Potentiate Local Antimelanoma Therapy.

Although nitric oxide (NO) has been emerging as a novel local anticancer agent because of its potent cytotoxic effects and lack of off-target side effects, its clinical applications remain a challenge because of the short effective diffusion distance of NO that limits its anticancer activity. In this study, we synthesized albumin-coated poly(lactic-co-glycolic acid) (PLGA)-conjugated linear polyethylenimine diazeniumdiolate (LP/NO) nanoparticles (Alb-PLP/NO NPs) that possess tumor-penetrating and NO-releasing properties for an effective local treatment of melanoma. Sufficient NO-loading and prolonged NO-releasing characteristics of Alb-PLP/NO NPs were acquired through PLGA-conjugated LP/NO copolymer (PLP/NO) synthesis, followed by nanoparticle fabrication. In addition, tumor penetration ability was rendered by the electrostatic adsorption of the albumin on the surface of the nanoparticles. The Alb-PLP/NO NPs showed enhanced intracellular NO delivery efficiency and cytotoxicity to B16F10 murine melanoma cells. In B16F10-tumor-bearing mice, the Alb-PLP/NO NPs showed improved extracellular matrix penetration and spatial distribution in the tumor tissue after intratumoral injection, resulting in enhanced antitumor activity. Taken together, the results suggest that Alb-PLP/NO NPs represent a promising new modality for the local treatment of melanoma.

Identifying Characteristics of Frailty in Female Mice Using a Phenotype Assessment Tool.

Preclinical studies are important in identifying the underlying mechanisms contributing to frailty. Frailty studies have mainly focused on male rodents with little directed at female rodents. Therefore, the purposes of this study were to identify the onset and prevalence of frailty across the life span in female mice, and to determine if frailty predicts mortality. Female C57BL/6 (n = 27) mice starting at 17 months of age were assessed across the life span using a frailty phenotype, which included body weight, walking speed, strength, endurance, and physical activity. The onset of frailty occurred at approximately 17 months (1/27 mice), with the prevalence of frailty increasing thereafter. At 17 months, 11.1% of the mice were pre-frail and by 26 months peaked at 36.9%. The percentage of frail mice progressively increased up to 66.7% at 32 months. Non-frail mice lived to 29 months whereas frail/pre-frail mice lived only to 26 months (p = .04). In closing, using a mouse frailty phenotype, we are able to identify that the prevalence of frailty in female mice increases across the life span and accurately predicts mortality. Together, this frailty phenotype has the potential to yield information about the underlying mechanisms contributing to frailty.

Short-Term ONX-0914 Administration: Performance and Muscle Phenotype in Mdx Mice.

Duchenne muscular dystrophy (DMD) is a severe muscle-wasting disease. Although the lack of dystrophin protein is the primary defect responsible for the development of DMD, secondary disease complications such as persistent inflammation contribute greatly to the pathogenesis and the time-dependent progression of muscle destruction. The immunoproteasome is a potential therapeutic target for conditions or diseases mechanistically linked to inflammation. In this study, we explored the possible effects of ONX-0914 administration, an inhibitor specific for the immunoproteasome subunit LMP7 (ß5i), on motor performance, muscular pathology and protein degradation in 7-week old MDX mice, an age when the dystrophic muscles show extensive degeneration and regeneration. ONX-0914 (10 mg/kg) was injected subcutaneously on Day 2, 4, and 6. The mice were evaluated for physical performance (walking speed and strength) on Day 1 and 8. We show that this short-term treatment of ONX-0914 in MDX mice did not alter strength nor walking speed. The physical performance findings were consistent with no change in muscle inflammatory infiltration, percentage of central nuclei and proteasome content. Taken together, muscle structure and function in the young adult MDX mouse model are not altered with ONX-0914 treatment, indicating the administration of ONX-0914 during this critical time period does not exhibit any detrimental effects and may be an effective treatment of secondary complications of muscular dystrophy after further investigations.

Enhanced Viability of Probiotics against Gastric Acid by One-Step Coating Process with Poly-L-Lysine: In Vitro and In Vivo Evaluation.

Due to their low acid tolerance, a majority of probiotics face diculties with regard tosurviving in the gastric environment long enough to reach the intestinal surfaces where they colonizeand provide health benefits. We prepared a probiotic delivery system that can enhance their viabilityin acidic conditions by developing a one-step poly-L-lysine (PLL) coating process. We determinedwhether the coating process was successful by measuring the zeta potential and observing it withconfocal scanning microscopy. PLL-coated L. plantarum (PLL-LP), incubated in a solution of pH 2 for2 h, exhibited a higher viability (6.86 0.12 log CFU/mL of viable cells) than non-coated L. plantarum(non-coated LP), which exhibited only 2.7 1.23 log CFU/mL of viable cells. In addition, a higheramount of L. plantarum was detected in the feces of mice orally administered PLL-LP (6.2 0.4 logCFU/g of feces) than in the feces of the control groups. In addition to enhancing probiotic viability inpH 2 solution, the PLL coating showed no eect on the probiotic growth pattern and the viability ofeither freeze-dried L. plantarum or L. plantarum, stored at ?20 C and 4 C, respectively. Overall, theseresults indicated that the PLL coating is a promising potential probiotic delivery system.

Nitric Oxide-Releasing S-Nitrosoglutathione-Conjugated Poly(Lactic-Co-Glycolic Acid) Nanoparticles for the Treatment of MRSA-Infected Cutaneous Wounds.

S-nitrosoglutathione (GSNO) has emerged as a potent agent for the treatment of infected cutaneous wounds. However, fabrication of GSNO-containing nanoparticles has been challenging due to its high hydrophilicity and degradability. The present study aimed to fabricate nanoparticles using newly synthesized GSNO-conjugated poly(lactic-co-glycolic acid) (PLGA) (GSNO-PLGA; GPNPs). Since hydrophilic GSNO was covalently bound to hydrophobic PLGA, loss of GSNO during the nanoparticle fabrication process was minimized, resulting in sufficient loading efficiency (2.32% of GSNO, 0.07 µmol/mg of NO). Real-time NO release analysis revealed biphasic NO release by GPNPs, including initial burst release within 3 min and continuous controlled release for up to 11.27 h, due to the differential degradation rates of the -SNO groups located at the surface and inside of GPNPs. Since GPNPs could deliver NO more efficiently than GSNO in response to increased interaction with bacteria, the former showed enhanced antibacterial effects against methicillin-resistant Staphylococcus aureus (MRSA) at the same equivalent concentrations of NO. Finally, the facilitating effects of GPNPs on infected wound healing were demonstrated in MRSA-challenged full-thickness wound mouse model. Collectively, the results suggested GPNPs as an ideal nanoparticle formulation for the treatment of MRSA-infected cutaneous wounds.