Modeling the Maturation of the Vocal Fold Lamina Propria Using a Bioorthogonally Tunable Hydrogel Platform.

Toward the goal of establishing an engineered model of the vocal fold lamina propria (LP), mesenchymal stem cells (MSCs) are encapsulated in hyaluronic acid (HA)-based hydrogels employing tetrazine ligation with strained alkenes. To mimic matrix stiffening during LP maturation, diffusion-controlled interfacial bioorthogonal crosslinking is carried out on the soft cellular construct using HA modified with a ferocious dienophile, trans-cyclooctene (TCO). Cultures are maintained in MSC growth media for 14 days to afford a model of a newborn LP that is homogeneously soft (nLP), a homogeneously stiffened construct zero (sLP0) or 7 days (sLP7) post cell encapsulation, and a mature LP model (mLP) with a stiff top layer and a soft bottom layer. Installation of additional HA crosslinks restricts cell spreading. Compared to the nLP controls, sLP7 conditions upregulate the expression of fibrous matrix proteins (Col I, DCN, and FN EDA), classic fibroblastic markers (TNC, FAP, and FSP1), and matrix remodeling enzymes (MMP2, TIMP1, and HAS3). Day 7 stiffening also upregulates the catabolic activities, enhances ECM turnover, and promotes YAP expression. Overall, in situ delayed matrix stiffening promotes a fibroblast transition from MSCs and enhances YAP-regulated mechanosensing.

Dynamic modulation of matrix adhesiveness induces epithelial-to-mesenchymal transition in prostate cancer cells in 3D.

Synthetic matrices with dynamic presentation of cell guidance cues are needed for the development of physiologically relevant in vitro tumor models. Towards the goal of mimicking prostate cancer progression and metastasis, we engineered a tunable hyaluronic acid-based hydrogel platform with protease degradable and cell adhesive properties employing bioorthogonal tetrazine ligation with strained alkenes. The synthetic matrix was first fabricated via a slow tetrazine-norbornene reaction, then temporally modified via a diffusion-controlled method using trans-cyclooctene, a fierce dienophile that reacts with tetrazine with an unusually fast rate. The encapsulated DU145 prostate cancer single cells spontaneously formed multicellular tumoroids after 7 days of culture. In situ modification of the synthetic matrix via covalent tagging of cell adhesive RGD peptide induced tumoroid decompaction and the development of cellular protrusions. RGD tagging did not compromise the overall cell viability, nor did it induce cell apoptosis. In response to increased matrix adhesiveness, DU145 cells dynamically loosen cell-cell adhesion and strengthen cell-matrix interactions to promote an invasive phenotype. Characterization of the 3D cultures by immunocytochemistry and gene expression analyses demonstrated that cells invaded into the matrix via a mesenchymal like migration, with upregulation of major mesenchymal markers, and down regulation of epithelial markers. The tumoroids formed cortactin positive invadopodia like structures, indicating active matrix remodeling. Overall, the engineered tumor model can be utilized to identify potential molecular targets and test pharmacological inhibitors, thereby accelerating the design of innovative strategies for cancer therapeutics.

Matrix Adhesiveness Regulates Myofibroblast Differentiation from Vocal Fold Fibroblasts in a Bio-orthogonally Cross-linked Hydrogel.

Repeated mechanical and chemical insults cause an irreversible alteration of extracellular matrix (ECM) composition and properties, giving rise to vocal fold scarring that is refractory to treatment. Although it is well known that fibroblast activation to myofibroblast is the key to the development of the pathology, the lack of a physiologically relevant in vitro model of vocal folds impedes mechanistic investigations on how ECM cues promote myofibroblast differentiation. Herein, we describe a bio-orthogonally cross-linked hydrogel platform that recapitulates the alteration of matrix adhesiveness due to enhanced fibronectin deposition when vocal fold wound healing is initiated. The synthetic ECM (sECM) was established via the cycloaddition reaction of tetrazine (Tz) with slow (norbornene, Nb)- and fast (trans-cyclooctene, TCO)-reacting dienophiles. The relatively slow Tz-Nb ligation allowed the establishment of the covalent hydrogel network for 3D cell encapsulation, while the rapid and efficient Tz-TCO reaction enabled precise conjugation of the cell-adhesive RGDSP peptide in the hydrogel network. To mimic the dynamic changes of ECM composition during wound healing, RGDSP was conjugated to cell-laden hydrogel constructs via a diffusion-controlled bioorthognal ligation method 3 days post encapsulation. At a low RGDSP concentration (0.2 mM), fibroblasts residing in the hydrogel remained quiescent when maintained in transforming growth factor beta 1 (TGF-ß1)-conditioned media. However, at a high concentration (2 mM), RGDSP potentiated TGF-ß1-induced myofibroblast differentiation, as evidenced by the formation of an actin cytoskeleton network, including F-actin and alpha-smooth muscle actin. The RGDSP-driven fibroblast activation to myofibroblast was accompanied with an increase in the expression of wound healing-related genes, the secretion of profibrotic cytokines, and matrix contraction required for tissue remodeling. This work represents the first step toward the establishment of a 3D hydrogel-based cellular model for studying myofibroblast differentiation in a defined niche associated with vocal fold scarring.

Nephrotic syndrome of minimal change disease following exposure to mercury-containing skin lightening cream: A case report and literature review.

Long-term exposure to mercury-containing skin lightening cream can cause mercury-related nephropathy, among which, membranous nephropathy (MN) and minimal change disease (MCD) are the main pathological types. In contrast to these two conditions, MCD with IgA deposition is not a common disease. In the present study, we report a 65-year-old Asian woman who developed nephrotic syndrome following long-term use of mercury-containing skin lightening cream. The urine mercury level of the patient was significantly increased, and the results of the renal biopsy indicated diagnosis of MCD with IgA deposition. Following three courses of treatment with sodium dimercaptopropane sulfonate (DMPS) alone and discontinuation of the skin cream, the symptoms of the patient were relieved without use of glucocorticoids, with proteinuria turning negative and a significant reduction in urine mercury levels. During the 6-month follow-up period, routine urinalysis remained normal. By reviewing relevant published literature, we summarized the pathological characteristics, possible mechanism of action, and treatment strategies of mercury poisoning-related MCD. The possibility of mercury poisoning should be considered for patients with nephropathy and history of use of skin lightening cosmetics. In these patients, the urine mercury levels should be measured in time so that mercury removal therapy can be implemented early.

Dioscin-loaded zein nanoparticles alleviate lipopolysaccharide-induced acute kidney injury via the microRNA-let 7i signalling pathways.

The present study investigates the potential role of dioscin (DIO) in the lipopolysaccharide (LPS)-induced kidney injury. For this purpose, DIO-loaded zein nanoparticles (DIO-ZNPs) were formulated and evaluated for physicochemical parameters. The DIO-ZNPs exhibited a controlled release of drug compared with that of the free drug suspension. Results showed that the cell viability of NRK-52E consistently decreased with the increase in LPS from 0.01 µg/ml to 2 µg/ml. When compared with LPS, DIO-induced NPs showed 1.10-, 1.32-, 1.57- and 1.92-fold increase in the cell viability for concentrations of 20 µg/ml, 50 µg/ml, 100 µg/ml and 200 µg/ml, respectively. DIO-ZNPs exhibited the most remarkable recovery in the cell proliferation compared with free DIO as shown by the cellular morphology analysis. Furthermore, Annexin-V staining analysis showed that the LPS-treated cells possess the lowest green fluorescence indicating fewer viable cells, whereas DIO-ZNPs exhibited the maximum green fluorescence comparable with that of the non-treated cells indicating maximum cell viability. Furthermore, the results show that DIO-ZNPs significantly increased the expression of miR-let-7i in the epithelial kidney cells, whereas the expression levels of TLR4 were significantly downregulated compared with that of the LPS-treated cells. In conclusion, miR-let-7i could be an interesting therapeutic target and nanoparticle-based DIO could be a potential candidate in the management of acute kidney injury.

To study the intervention mechanism of pediatric massage on intestinal flora and host metabolism in children with anorexia.

BACKGROUND: As a common and frequent disease in pediatric patients, pediatric anorexia (PN) poses a serious threat to childhood growth and health. In recent years, societal changes in lifestyle and diet have increased the incidence of this PN, which has attracted extensive attention from both the medical community and parents. It has been shown that massage therapy represents an effective intervention for the treatment of anorexia, but investigation on its mechanism(s) of action remains limited. In this study, we will explore the biological mechanism(s) of PN from the perspective of intestinal flora, to further reveal its site of action and therapeutic mechanism(s). METHODS: A total of 60 healthy children will be randomly selected for physical examination. According to a random number generated by a computer, children with anorexia who meet the inclusion criteria will be selected. In strict accordance with the time sequence of inclusion, subjects will be randomly assigned to either the massage or control group (n = 60 per group). The blank group will receive no treatment. Children in the massage group will receive a designated massage protocol. The control group will be administered oral Jianweixiaoshi tablets over 4 weeks. Each group will be compared for intestinal flora structure, fecal short chain fatty acids levels, serum trace elements, urine D-xylose-excretion rates, gastric fluid emptying, gastric motility, and hemoglobin levels before and after treatment. RESULTS: We will review the clinical trial registry in China (http://www.chictr.org.cn/searchprojen.aspx), peer-reviewed journals and academic conferences. CONCLUSION: This study will verify the intervention mechanism(s) of pediatric massage on intestinal flora and host metabolism in children with anorexia. TRIAL REGISTRATION NUMBER: ChiCTR2000033274.

Enantioseparation using chitosan 2-isopropylthiourea-3,6-dicarbamate derivatives as chiral stationary phases for high-performance liquid chromatography.

A new class of chitosan derivatives with an isopropylthiourea at the 2-position and various carbamates at the 3,6-positions of the glucosamine skeleton was synthesized by the selective thiocarbamoylation of the 2-amino group. The chiral stationary phases (CSPs) were then prepared by coating the obtained chitosan 2-isopropylthiourea-3,6-dicarbamate derivatives onto silica gel. The enantioseparation property of the chitosan-based CSPs was assessed with twelve racemates by high-performance liquid chromatography (HPLC). The CSPs displayed a characteristic enantioseparation power, which seemed to be significantly affected by the 3,6-substituents of the glucosamine unit. The chitosan derivatives with the 3,6-diphenylcarbamate, except for 2-methylphenylcarbamate, possessed higher enantioseparation abilities than those with the 3,6-dicyclohexylcarbamate. Compared to other chitosan derivatives with 2-various substituents and commercialized Chiralcel OD, the chitosan 2-isopropylthiourea derivatives revealed a relatively higher enantioselectivity for some racemic compounds.

The association between expression of IFIT1 in podocytes of MRL/lpr mice and the renal pathological changes it causes: An animal study.

Renal damage is the major cause of SLE associated mortality, and IFIT1expression was elevated in SLE cases in accordance of previous studies. Therefore, we conducted an animal study to identify the role of IFIT1 expression in renal pathological changes.18 female MRL/lpr mice and same number of female BALB/c mice were enrolled in present study. Quantitative analysis of urine protein, Complement C3 and C4, and anti-ds DNA antibody were conducted. HE and PAS staining and TEM analysis were employed to observe the pathological changes in renal tissue. Significant elevation on urine protein and anti-dsDNA and reduction on Complement C3 and C4 were observed in MRL/lpr mice when comparing the controls in same age. Staining and TEM analysis observed several pathological changes in glomerulus among MRL/lpr mice, including cellular enlargement, basement membrane thickening, and increased cellularcasts. The linear regression analysis found the optical density of IFIT1 was inversely associated with F-actin, Nephrin, and Podocin, but not Synatopodin. In summary, IFIT1 expression is associated with podocytes damage, and capable of suppressing some proteins essential to glomerular filtration.

[Treatment of 60 cases of allergic rhinitis mainly with point-through-point method].

OBJECTIVE: To search for an effective therapy for allergic rhinitis. METHODS: One hundred and twenty cases of allergic rhinitis were randomly divided into a treatment group (n=60) treated mainly with penetration needling, Yintang (EX-HN 3)-through-Bigen (root of nose), Sibai (ST 2)-through-Bigen, Yingxiang (LI 20)-through-Bigen, and a control group (n=60) treated with oral Biyankang, 4 tablets each time, 3 times daily. Their therapeutic effects were compared. RESULTS: The total effective rate was 85.0% in the treatment group and 60.0% in the control group, with a very significant difference between the two groups (P<0.01). CONCLUSION: The penetration needling as main therapy has a definite therapeutic effect on allergic rhinitis, which is significantly better than that of oral Biyankang.