Application of Milk Exosomes for Musculoskeletal Health: Talking Points in Recent Outcomes.

Milk is a nutrient-rich food source, and among the various milks, breast milk is a nutrient source provided by mothers to newborns in many mammals. Exosomes are nano-sized membranous extracellular vesicles that play important roles in cell-to-cell communication. Exosomes originate from endogenous synthesis and dietary sources such as milk. Discovered through electron microscopy as floating vesicles, the existence of exosomes in human milk was confirmed owing to a density between 1.10 and 1.18 g/mL in a sucrose gradient corresponding to the known density of exosomes and detection of MHC classes I and II, CD63, CD81, and CD86 on the vesicles. To date, milk exosomes have been used for treating many diseases, including cancers, and are widely proposed as promising carriers for the delivery of chemotherapeutic agents. However, few studies on milk exosomes focus on geriatric health, especially sarcopenia and osteoporosis related to bone and muscle. Therefore, the present study focused on milk exosomes and their cargoes, which are potential candidates for dietary supplements, and when combined with drugs, they can be effective in treating musculoskeletal diseases. In this review, we introduce the basic concepts, including the definition, various sources, and cargoes of milk exosomes, and exosome isolation and characterization methods. Additionally, we review recent literature on the musculoskeletal system and milk exosomes. Since inflammation and oxidative stress underly musculoskeletal disorders, studies reporting the antioxidant and anti-inflammatory properties of milk exosomes are also summarized. Finally, the therapeutic potential of milk exosomes in targeting muscle and bone health is proposed.

Protective Role of Ethanol Extract of Cibotium barometz (Cibotium Rhizome) against Dexamethasone-Induced Muscle Atrophy in C2C12 Myotubes.

Sarcopenia is a progressive muscle disease characterized by the loss of skeletal muscle mass, strength, function, and physical performance. Since the disease code was assigned, attention has been focused on natural products that can protect against muscle atrophy. Cibotium barometz (Cibotium Rhizome) has been used as an herbal medicine for the treatment of bone or joint diseases in Asian countries. However, no studies have identified the mechanism of action of Cibotium Rhizome on muscle atrophy related to sarcopenia at the site of myotubes. The aim of this study was to investigate the improvement effect of the ethanol extract of Cibotium Rhizome (ECR) on dexamethasone-induced muscle atrophy in an in vitro cell model, i.e., the C2C12 myotubes. High-performance liquid chromatography was performed to examine the phytochemicals in ECR. Seven peaks in the ECR were identified, corresponding to the following compounds: protocatechuic acid, (+)-catechin hydrate, p-coumaric acid, ellagic acid, chlorogenic acid, caffeic acid, and ferulic acid. In atrophy-like conditions induced by 100 µM dexamethasone for 24 h in C2C12, ECR increased the expression of the myosin heavy chain, p-Akt, the p-mammalian target of rapamycin (mTOR), p-p70S6K, and repressed the expression of regulated in development and DNA damage responses 1 (REDD1), kruppel-like factor 15 (KLF 15), muscle atrophy F-box, and muscle-specific RING finger protein-1 in C2C12. In addition, ECR alleviated dexamethasone-induced muscle atrophy by repressing REDD1 and KLF15 transcription in C2C12 myotubes, indicating the need for further studies to provide a scientific basis for the development of useful therapeutic agents using ECR to alleviate the effects of skeletal muscle atrophy or sarcopenia.

Effects of Allium macrostemon Bunge Extract on Adipose Tissue Inflammation and Hepatic Endoplasmic Reticulum Stress in High-Fat Diet-Fed and Bisphenol A-Treated C57BL/6N Mice.

The simultaneous exposure to a high-fat (HF) diet and to bisphenol A (BPA) from delivered foods and food-delivery containers is on the rise in humans, according to the increased frequency of food delivery during the COVID-19 pandemic. This co-exposure could cause harmful tissue toxicity in the human body. Here, the preventive effect of Allium macrostemon Bunge (AM) extract against dysfunction in adipose tissue and the liver under co-exposure to BPA and an HF diet was examined using mice. C57BL/6N mice were divided into four groups (n = 6 or 7/group) according to diet and treatment: control diet with vehicle (CON), HF diet with vehicle (HF), HF diet with an oral injection of BPA (HF + BP), and HF diet with an oral injection of BPA and AM extract (HF + BP + AM). HF feeding increased body weight gain compared to CON feeding, while BP + HF and BP + HF + AM feeding suppressed body weight gain compared with HF feeding. The BP + HF group had lower body weight than the HF group, but the two groups had similar epididymal fat mass. The HF + BP + AM group showed lower pro-inflammatory gene expression levels in adipose tissue and epididymal fat mass compared to the HF + BP group. Altered endoplasmic reticulum (ER) stress response in the liver was partly observed in the HF + BP group, as shown by increased total phosphorylated Jun N-terminal kinase protein levels compared to those in the HF group. In addition, ecdysterone 25-O-ß-D-glucopyranoside and 6-gingerol were identified in AM extract by mass spectrometry and molecular networking analysis. In summary, the AM extract diminished adipose tissue inflammation and hepatic ER stress in an HF diet and BPA co-exposure condition. To utilize AM as a potential food component to alleviate the harmful effect of an HF diet and BPA exposure, further research investigating the specific impact of AM extract supplementation using additional experimental groups or various treatment doses is warranted.

Nonclinical safety assessment and immunogenicity of rVSVInd(GML)-mspSGtc vaccine for SARS-CoV-2 in rabbits.

The worldwide health, economic, and societal consequences of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic have been devastating. The primary strategy to prevent new infectious diseases is to vaccinate the majority of people worldwide. However, the significant hurdles that are faced include vaccine safety concerns and vaccine reluctance. Among the various types of vaccines, the recombinant vesicular stomatitis virus (rVSV) is a promising candidate owing to its safety and efficacy. Therefore, we investigated the toxicity, immunogenicity, and local tolerance of the rVSVInd(GML)-mspSGtc vaccine against SARS-CoV-2. New Zealand White (NZW) rabbits were administered single or three repeated intramuscular injections of rVSVInd(GML)-mspSGtc every 2 weeks, followed by a 4-week recovery period. Male and female rabbits were randomly assigned into three groups: a control group and two dose-level groups (1 × 109 and 4 × 109 PFU/mL). Treatment-related changes included a temporary increase in body temperature and local inflammation at the injection site. These findings indicated recovery or a trend toward recovery, with no overt systemic toxicity. Immunogenicity analysis results suggested that rVSVInd(GML)-mspSGtc elicited a robust dose-dependent immune response in terms of neutralizing antibodies and IgG antibodies against the SARS-CoV-2 spike protein. In addition, the immune response intensity was increased by repeated vaccine administration. In conclusion, both the approximate lethal dose and the no observed adverse effect level for rVSVInd(GML)-mspSGtc exceeded 4 × 109 PFU/mL in NZW rabbits. Overall, rVSVInd(GML)-mspSGtc induced no adverse effects at the maximum dosage tested; however, its efficacy warrants further clinical evaluation.

Correction: A vesicular stomatitis virus-based prime-boost vaccination strategy induces potent and protective neutralizing antibodies against SARS-CoV-2.

[This corrects the article DOI: 10.1371/journal.ppat.1010092.].

Association of Micronutrients and Handgrip Strength in Korean Older Population: A Cross-Sectional Study.

Sarcopenia is characterized by the loss of skeletal muscle mass, strength, and physical performance. Dynapenia and kratopenia are described as the loss of muscle strength and power. Nutritional intake status is one of the factors affecting the prevention of an age-related muscle decline such as sarcopenia, dynapenia, or kratopenia in older populations. This study aimed to investigate the association between the intake of micronutrients and handgrip strength in 1254 individuals (546 men and 708 women) of the Korean older population from the most recent dataset. They were analyzed and divided into two groups: a LHS group with low handgrip strength (<28 kg for men and <18 kg for women) and a normal group with normal handgrip strength. Logistic regression analysis was performed to estimate the odds ratios (ORs) and 95% confidence intervals (Cis) of the associations between micronutrient intakes and low handgrip strength in Korean older population by gender. Among micronutrients, insufficient potassium intake showed a significant association with low handgrip strength for men (OR: 3.159, 95% CI: 1.164−8.578) and women (OR: 2.793, 95% CI: 1.380−5.654) aged ≥65 years, respectively (p = 0.005 for men, p = 0.024 for women), as a result of adjusting for all confounding factors that could affect low handgrip strength. In conclusion, potassium intake among micronutrients in Korean older populations with low handgrip strength might need continuous monitoring for the intervention or prevention of dynapenia or sarcopenia.

A vesicular stomatitis virus-based prime-boost vaccination strategy induces potent and protective neutralizing antibodies against SARS-CoV-2.

The development of safe and effective vaccines to prevent SARS-CoV-2 infections remains an urgent priority worldwide. We have used a recombinant vesicular stomatitis virus (rVSV)-based prime-boost immunization strategy to develop an effective COVID-19 vaccine candidate. We have constructed VSV genomes carrying exogenous genes resulting in the production of avirulent rVSV carrying the full-length spike protein (SF), the S1 subunit, or the receptor-binding domain (RBD) plus envelope (E) protein of SARS-CoV-2. Adding the honeybee melittin signal peptide (msp) to the N-terminus enhanced the protein expression, and adding the VSV G protein transmembrane domain and the cytoplasmic tail (Gtc) enhanced protein incorporation into pseudotype VSV. All rVSVs expressed three different forms of SARS-CoV-2 spike proteins, but chimeras with VSV-Gtc demonstrated the highest rVSV-associated expression. In immunized mice, rVSV with chimeric S protein-Gtc derivatives induced the highest level of potent neutralizing antibodies and T cell responses, and rVSV harboring the full-length msp-SF-Gtc proved to be the superior immunogen. More importantly, rVSV-msp-SF-Gtc vaccinated animals were completely protected from a subsequent SARS-CoV-2 challenge. Overall, we have developed an efficient strategy to induce a protective response in SARS-CoV-2 challenged immunized mice. Vaccination with our rVSV-based vector may be an effective solution in the global fight against COVID-19.

Induction of protective immune responses against a lethal Zika virus challenge post-vaccination with a dual serotype of recombinant vesicular stomatitis virus carrying the genetically modified Zika virus E protein gene.

The development of a vaccine to prevent Zika virus (ZIKV) infection has been one of the priorities in infectious disease research in recent years. There have been numerous attempts to develop an effective vaccine against ZIKV. It is imperative to choose the safest and the most effective ZIKV vaccine from all candidate vaccines to control this infection globally. We have employed a dual serotype of prime-boost recombinant vesicular stomatitis virus (VSV) vaccine strategy, to develop a ZIKV vaccine candidate, using a type 1 IFN-receptor knock-out (Ifnar-/-) mouse model for challenge studies. Prime vaccination with an attenuated recombinant VSV Indiana serotype (rVSVInd) carrying a genetically modified ZIKV envelope (E) protein gene followed by boost vaccination with attenuated recombinant VSV New Jersey serotype (rVSVNJ) carrying the same E gene induced robust adaptive immune responses. In particular, rVSV carrying the ZIKV E gene with the honeybee melittin signal peptide (msp) at the N terminus and VSV G protein transmembrane domain and cytoplasmic tail (Gtc) at the C terminus of the E gene induced strong protective immune responses. This vaccine regimen induced highly potent neutralizing antibodies and T cell responses in the absence of an adjuvant and protected Ifnar-/- mice from a lethal dose of the ZIKV challenge.

Fermented Garlic Extract Increases Oxygen Consumption and UCP-1 mRNA Expression in Human Adipose-Derived Stem Cells.

Fermented garlic, often called black garlic, is a traditional food ingredient used in Asian cuisine and possesses various health benefits including anti-obesity activity. The anti-obesity effects of fermented garlic might, in part, might be mediated through direct actions of its components on adipocytes. To test this hypothesis, we examined whether fermented garlic extract might stimulate the metabolic activity of human adipose-derived stem cells (ADSCs) in culture. Cell viability measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium (MTT) assay exhibited a complex doseresponse relationship. The lowest concentration (0.4 mg/ml) reduced cell viability (P<0.05 compared to no extract, Bonferroni's multiple comparison), whereas higher concentrations (0.8 and 1.0 mg/ml) resulted in higher cell viability (P<0.05 as compared to 0.4 mg/ml). However, the extract at concentrations >2 mg/ml markedly decreased cell viability. Higher cell viability observed following treatment with 0.8~1.0 mg/ml might be associated with raised oxygen consumption. Fluorescent dye-based measurement revealed that the garlic extract at 1.0 mg/ml significantly increased oxygen consumption. We also detected a significant increase in mRNA expression levels of uncoupling protein-1 (UCP- 1). These findings suggest that fermented garlic stimulates the basal metabolic activity of human ADSCs.

Molecular characterization of glucosinolates and carotenoid biosynthetic genes in Chinese cabbage (Brassica rapa L. ssp. pekinensis).

The present study aimed to investigate the contents of glucosinolates (GSLs) and carotenoids in eleven varieties of Chinese cabbage in relation to the expression level of the important transcription factors. MS and HPLC analysis identified the presence of 13 GSLs (progoitrin, sinigrin, glucoalyssin, gluconapoleiferin, gluconapin, glucocochlearin, glucobrassicanapin, glucoerucin, 4-hydroxyglucobrassicin, glucobrassicin, 4-methoxyglucobrassicin, neoglucobrassicin and gluconasturtiin) and four carotenoids (lutein, zeaxanthin, α-carotene and ß-carotene). GSL contents were varied among the different cabbage varieties. The total GSL content ranged from 2.7 to 57.88 µmol/g DW. The proportion of gluconapin (54%) and glucobrassicanapin (22%) was higher in all the varieties, respectively. Results documented the variation in total and individual carotenoid contents that have also been observed among different varieties; however, the total carotenoid contents ranged from 289.12 to 1001.41 mg kg-1 DW (mean 467.66). Interestingly, the proportion of lutein (66.5) and ß-carotene (25.9) were higher than α-carotene (5.1) and zeaxanthin (2.5%). Consequently, the expression level of the regulatory gene, MYB28 was higher in 'K0648' and was directly proportional to GSL content. Similarly, the expression levels of 1-PSY were higher in 'K0112'; however, the expression levels of 2-ZDS, 3-LCYB, 4-LCYE, 5-CHXB and 7-NCED genes showed no significant difference. In addition, the correlation between GSL and carotenoid contents and gene expression level showed moderate significant difference in each Chinese cabbage.

Liver Histological Improvement After Administration of High-Dose Vitamin C in Guinea Pig with Nonalcoholic Steatohepatitis.

Background: Vitamin C is a strong antioxidant, and the health effects of vitamin C megadoses have not been validated despite the apparent health benefits. Therefore, the present study sought to confirm the effects of vitamin C megadoses. Materials and Methods : Four groups of six guinea pigs were used. Each group was fed one of the following diets for three weeks: normal diet, methionine choline-deficient diet, methionine choline-deficient diet + vitamin C megadose (MCD + vit C 2.5 g/kg/day), and methionine-choline deficient diet + ursodeoxycholic acid (MCD + UDCA 30 mg/kg/day). The MCD diet was given to induce nonalcoholic steatohepatitis, and UDCA was used to treat nonalcoholic steatohepatitis. Three weeks after initial diet administration, the results of biochemical tests and liver biopsy were compared between the groups. Results: The cytoplasm state was similar in the MCD + vit C and MCD + UDCA groups, exhibiting clearing of the cytoplasm and ballooning degeneration. However, macrovesicular steatosis was not observed in the MCD + vit C group. Aspartate transaminase and alanine transaminase were elevated significantly following vitamin C administration. Conclusions: The present study confirmed that alone vitamin C megadoses are potential remedies for nonalcoholic steatohepatitis, based on the liver biopsy results of guinea pigs that were unable to synthesize vitamin C.

Ghost probiotics with a combined regimen: a novel therapeutic approach against the Zika virus, an emerging world threat.

The Zika virus (ZIKV) used to be an obscure flavivirus closely related to dengue virus (DENV). Transmission of this epidemic pathogen occurs mainly via mosquitoes, but it is also capable of placental and sexual transmission. Although the characteristics of these viruses are well defined, infections are unpredictable in terms of disease severity, unusual clinical manifestations, unexpected methods of transmission, long-term persistence, and the development of new strains. Recently, ZIKV has gained huge medical attention following the large-scale epidemics around the world, and reported cases of congenital abnormalities associated with Zika virus infections which have created a public health emergency of international concern. Despite continuous research on ZIKV, no specific treatment or vaccine has been developed, excepting a preventive strategy for congenital ZIKV infection. Probiotics, known as GRAS, are bacteria that confer various health beneficial effects, and have been shown to be effective at curing a number of viral diseases by modulating the immune system. Furthermore, probiotic preparations consisting of dead cells and cellular metabolites, so-called "Ghost probiotics", can also act as biological response modifiers. Here, we review available information on the epidemiology, transmission, and clinical features of ZIKV, and on treatment and prevention strategies. In addition, we emphasize the use of probiotics and plant-based natural remedies and describe their action mechanisms, and the green technologies for microbial conversion, which could contribute to the development of novel therapies that may reduce the pathogenicity of ZIKV. Accordingly, we draw attention to new findings, unanswered questions, unresolved issues, and controversies regarding ZIKV.

Variation of quercetin glycoside derivatives in three onion (Allium cepa L.) varieties.

The aim of this study was to quantify the contents of individual quercetin glycosides in red, yellow and chartreuse onion by High Performance Liquid Chromatography (HPLC) analysis. Acid hydrolysis of individual quercetin glycosides using 6 M hydrochloric acid guided to identify and separate quercetin 7,4'-diglucoside, quercetin 3-glucoside, quercetin 4'-glucoside, and quercetin. The contents of total quercetin glycosides varied extensively among three varieties (ranged from 16.10 to 103.93 mg/g DW). Quercetin was the predominant compound that accounted mean 32.21 mg/g DW in red onion (43.6% of the total) and 127.92 mg/g DW in chartreuse onion (78.3% of the total) followed by quercetin 3-glucoside (28.83 and 24.16 mg/g DW) respectively. Quercetin 3-glucoside levels were much higher in yellow onion (43.85 mg/g DW) followed by quercetin 30.08 mg/g DW. Quercetin 4'-glucoside documented the lowest amount that documented mean 2.4% of the total glycosides. The varied contents of glycosides present in the different onion varieties were significant.

Antitumor and Immunomodulatory Effect of Gastrodia elata on Colon Cancer In Vitro and In Vivo.

Gastrodia elata Blume (GE) is a well-known kind of herb that has been used in traditional medicine for thousands of years. The extrusion of raw materials from it could improve flavor and enhance bioavailability in food and drug development. The purpose of this study is to investigate antitumor and immune boosting effects of extruded GE in human colon carcinoma cells, splenocytes, and mice-bearing CT26 colon carcinoma cell. Treatment with 100[Formula: see text][Formula: see text]g/mL of extruded GE decreased cell viability and induced the expression of Caspase-3 and Bax in HT29 cells ([Formula: see text]). When we performed DAPI staining, apoptotic bodies with condensed chromatin and fragmented nuclei, known as indicative of apoptotic morphology, increased 24[Formula: see text]h after treatment with 100[Formula: see text][Formula: see text]g/mL of extruded GE. Treatments with extruded GE significantly promoted splenocyte proliferation and IL-2 or IFN-[Formula: see text] secretion, compared with that of control cells ([Formula: see text]). The administration of extruded GE of 200 mg/kg/day decreased tumor growth and Ki-67 or [Formula: see text]-catenin expression in mice ([Formula: see text]). Additionally, we investigated the contents of compounds in extruded GE extracts using ultra performance liquid chromatography. The contents of p-hydroxylbenzyl alcohol and p-hydroxybenzaldehyde in extruded GE were 2.97[Formula: see text]mg/g and 0.04[Formula: see text]mg/g, respectively. It was supposed that antitumor and immunomodulatory effects of extruded GE might exert by the p-hydroxylbenzyl alcohol and p-hydroxybenzaldehyde of many compositions analyzed from extruded GE. These results suggest that extruded GE have the potential to be developed into a natural pharmaceutical and functional food as a cancer chemopreventive agent.

Variation of glucosinolates on position orders of flower buds in turnip rape (Brassica rapa).

To glucosinolate (GSL) contents on flower buds depending on their position orders in turnip rape (Brassica rapa), three Japanese 'Nabana' cultivars such as cv. No. 21 (Brassica rapa, early type), cv. Husanohana (B. rapa, late type) and cv. Norin No. 20 (B. napus) were investigated using HPLC analysis. Ten GSLs including glucoraphanin, sinigrin, glucoalyssin, napoleiferin, gluconapin, 4-hydroxyglucobrassicin, glucobrassicanapin, glucobrassicin, and gluconasturtiin were detected. Differences in individual and total GSL contents were found between two plant varieties, which are also depending on various developmental stages. Among the GSLs, gluconapin (mean 23.11 µmole/g dry weight (DW) and glucobrassicanapin (mean 13.41 µmole/g DW) documented the most abundant compounds and contributed average 39 and 27% of the total GSLs, but indolyl and aromatic GSLs together accounted >10% of the total GSLs. The presence of significant quantities of gluconapin in the cultivars should be studied more extensively, since the GSL is mainly responsible for the bitter taste.

Anti-inflammatory Potential of Quercetin-3-O-ß-D-("2"-galloyl)-glucopyranoside and Quercetin Isolated from Diospyros kaki calyx via Suppression of MAP Signaling Molecules in LPS-induced RAW 264.7 Macrophages.

Diospyros kaki (DK) contains an abundance of flavonoids and has been used in folk medicine in Korea for centuries. Here, we report for the first time the anti-inflammatory activities of Quercetin (QCT) and Quercetin 3-O-ß-("2"-galloyl)-glucopyranoside (Q32G) isolated from DK. We have determine the no cytotoxicity of Q32G and QCT against RAW 264.7 cells up to concentration of 50 µM. QCT and Q32G demonstrated potent anti-inflammatory activities by reducing expression of nitric oxide (NO), tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, IL-6 inducible NO synthase (iNOS), cyclooxygenase (COX)-2, and mitogen-activated protein kinase (MAPKs) in mouse RAW 264.7 macrophages activated with lipopolysaccharide (LPS). Both QCT or Q32G could decrease cellular protein levels of COX-2 and iNOS as well as secreted protein levels of NO, PGE2 , and cytokines (TNF-α, IL-1ß, and IL-6) in culture medium of LPS-stimulated RAW 264.7 macrophages. Immunoblot analysis showed that QCT and Q32G suppressed LPS-induced MAP kinase pathway proteins p-p38, ERK, and JNK. This study revealed that QCT and Q32G have anti-inflammatory potential, however Q32G possess comparable activity as that of QCT and could be use as adjuvant to treat inflammatory diseases.

Antiviral potential of a diterpenoid compound sugiol from Metasequoia glyptostroboides.

This research reports first time antiviral activity of sugiol, a diterpenoid isolated from Metasequoia glyptostroboides in terms of its ability to inhibit in vitro growth of H1N1 influenza virus. Antiviral potential of sugiol was evaluated through hcytopathogenic reduction assay using Madin-Darby canine kidney (MDCK) cell line. Sugiol (500 µg/ml) was found to exhibit considerable anti-cytopathic effect on MDCK cell line confirming its antiviral efficacy against H1N1 influenza virus. These findings strongly reinforce the suggestion that sugiol could be a candidate of choice in combinational regimen with potential antiviral efficacy.

Protective effect of heat-treated cucumber (Cucumis sativus L.) juice on alcohol detoxification in experimental rats.

In this study, heat-treated cucumber juice was assessed for its protective effect on blood alcohol levels and hepatic alcohol metabolic enzyme system in experimental rats. Initially, during detoxification of alcohol, all groups were orally dosed to 22% alcohol (6ml/kg body weight) along with different concentrations of heat-treated cucumber juice (10, 100 and 500mg/kg) and commercial goods for hangover-removal on sale (2ml/kg). Cucumber juice was dosed before 30 min, and simultaneously after 30min of alcohol administration, and its hepatoprotective effect on blood alcohol levels and hepatic alcohol metabolic enzyme system in experimental rats was evaluated. As a result, after 7h, remarkable reduction was found in the blood alcohol levels for all concentrations of cucumber juice treatment. Treatment with cucumber juice resulted in increasing dehydrogenase (ADH) and acetaldehyde dehydrogenase (ALDH) enzymatic activities in rat liver at 9h after alcohol administration thereby stimulated blood alcohol metabolism as compared with control group. The effect of heat-treated cucumber juice on alcohol detoxification was observed only in the rats treated before 30min from alcohol administration. These findings indicate that heat-treated cucumber juice has significant protective effect on alcohol detoxification in experimental rats, suggesting its usefulness in the treatment of liver injury caused by alcohol consumption.

Post-Transcriptional Regulation of the GASC1 Oncogene with Active Tumor-Targeted siRNA-Nanoparticles.

Basal-like breast cancer (BLBC) accounts for the most aggressive types of breast cancer, marked by high rates of relapse and poor prognoses and with no effective clinical therapy yet. Therefore, investigation of new targets and treatment strategies is more than necessary. Here, we identified a receptor that can be targeted in BLBC for efficient and specific siRNA mediated gene knockdown of therapeutically relevant genes such as the histone demethylase GASC1, which is involved in multiple signaling pathways leading to tumorigenesis. Breast cancer and healthy breast cell lines were compared regarding transferrin receptor (TfR) expression via flow cytometry and transferrin binding assays. Nanobioconjugates made of low molecular weight polyethylenimine (LMW-PEI) and transferrin (Tf) were synthesized to contain a bioreducible disulfide bond. siRNA complexation was characterized by condensation assays and dynamic light scattering. Cytotoxicity, transfection efficiency, and the targeting specificity of the conjugates were investigated in TfR positive and negative healthy breast and breast cancer cell lines by flow cytometry, confocal microscopy, RT-PCR, and Western blot. Breast cancer cell lines revealed a significantly higher TfR expression than healthy breast cells. The conjugates efficiently condensed siRNA into particles with 45 nm size at low polymer concentrations, showed no apparent toxicity on different breast cancer cell lines, and had significantly greater transfection and gene knockdown activity on mRNA and protein levels than PEI/siRNA leading to targeted and therapeutic growth inhibition post GASC1 knockdown. The synthesized nanobioconjugates improved the efficiency of gene transfer and targeting specificity in transferrin receptor positive cells but not in cells with basal receptor expression. Therefore, these materials in combination with our newly identified siRNA sequences are promising candidates for therapeutic targeting of hard-to-treat BLBC and are currently further investigated regarding in vivo targeting efficacy and biocompatibility.