Encapsulation of a novel peptide derived from histatin-1 in liposomes against initial enamel caries in vitro and in vivo.

OBJECTIVES: Biomimetic mineralization mediated by proteins and peptides is a promising strategy for enamel repair, and its specific application model needs more research. In this work, we exploited a liposomal delivery system for a novel peptide (DK5) derived from histatin-1 (DK5-Lips) as a new biomimetic mineralization strategy against initial enamel caries. MATERIALS AND METHODS: The DK5-Lips was prepared using calcium acetate gradient method and then the in vitro release, salivary stability, and cytotoxicity were studied. Initial enamel caries was created in bovine enamel blocks and subjected to pH-cycling model treated with DK5-Lips. Surface microhardness testing, polarized light microscopy (PLM), and transverse microradiography (TMR) were analyzed. Then the biocompatibility of DK5-Lips was evaluated in the caries model of Sprague-Dawley rats, and the anti-caries effect was assessed using Micro-CT analysis, Keyes scores, and PLM in vivo. RESULTS: DK5-Lips provided a mean particle size of (97.63 ± 4.94)nm and encapsulation efficiency of (61.46 ± 1.44)%, exhibiting a sustained release profile, excellent stability in saliva, and no significant toxicity on human gingival fibroblasts (HGFs). The DK5-Lips group had higher surface microhardness recovery, shallower caries depth, and less mineral loss in bovine enamel. Animal experiments showed higher volume and density values of residual molar enamel, lower Keyes score, and shallower lesion depth of the DK5-Lips group with good biocompatibility. CONCLUSION: As a safe and effective application model, DK5-Lips could significantly promote the remineralization of initial enamel caries both in vitro and in vivo. CLINICAL RELEVANCE: The potential of liposome utilization as vehicle for oral delivery of functional peptides may provide a new way for enamel restoration.

TVH-19, a synthetic peptide, induces mineralization of dental pulp cells in vitro and formation of tertiary dentin in vivo.

Functional peptides derived from the active domains of odontogenesis-related proteins have been reported to promote dental hard tissue regeneration. The purpose of this study was to evaluate the effects of an artificially synthesized peptide, TVH-19, on odontoblast differentiation and tertiary dentin formation in indirect pulp capping (IPC) using in vitro and in vivo experiments. TVH-19 did not exhibit any effect on the proliferation of human dental pulp cells (hDPCs) but significantly promoted cell migration, compared with the control (p < 0.05). TVH-19-treated hDPCs showed significantly higher alkaline phosphatase (ALP) activity and stronger alizarin red staining (ARS) reactivity than the control group (p < 0.05). TVH-19 also upregulated the mRNA and protein expression levels of odontogenic genes. After generating IPC in rats, the samples of teeth were studied using micro-computed tomography (Micro-CT), hematoxylin & eosin (HE) staining, and immunohistochemical staining to investigate the functions of TVH-19. The in vivo results showed that TVH-19 induced the formation of tertiary dentin, and reduced inflammation and apoptosis, as evident from the downregulated expression of interleukin 6 (IL-6) and cleaved-Caspase-3 (CL-CASP3). Overall, the results of our study suggest that TVH-19 induces differentiation of hDPCs, promotes tertiary dentin formation, relieves inflammation, and reduces apoptosis, indicating the potential applications of TVH-19 in IPC.

Development and validation a simple model for identify malignant ascites.

The differential diagnosis of benign ascites and malignant ascites is incredibly challenging for clinicians. This research aimed to develop a user-friendly predictive model to discriminate malignant ascites from non-malignant ascites through easy-to-obtain clinical parameters. All patients with new-onset ascites fluid were recruited from January 2014 to December 2018. The medical records of 317 patients with ascites for various reasons in Renmin Hospital of Wuhan University were collected and reviewed retrospectively. Thirty-six parameters were included and selected using univariate logistic regression, multivariate logistic regression, and receiver operating characteristic (ROC) curve analyses to establish a mathematical model for differential diagnosis, and its diagnostic performance was validated in the other groups. Age, cholesterol, hypersensitivity C-reactive protein (hs-CRP) in serum, ascitic fluid adenosine deaminase (AF ADA), ascitic fluid lactate dehydrogenase (AF LDH) involvement in a 5-marker model. With a cut-off level of 0.83, the sensitivity, specificity, accuracy, and area under the ROC of the model for identifying malignant ascites in the development dataset were 84.7%, 88.8%, 87.6%, and 0.874 (95% confidence interval [CI], 0.822-0.926), respectively, and 80.9%, 82.6%, 81.5%, and 0.863 (95% CI,0.817-0.913) in the validation dataset, respectively. The diagnostic model has a similar high diagnostic performance in both the development and validation datasets. The mathematical diagnostic model based on the five markers is a user-friendly method to differentiate malignant ascites from benign ascites with high efficiency.

[Twin girls infected with SARS-CoV-2].

This article reports the diagnosis and treatment of twin girls who were diagnosed with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in Hunan Province, China. The twin girls, aged 1 year and 2 months, were admitted on January 29, 2020 due to fever for one day and cough and sneezing for two days respectively. Both recovered after symptomatic treatment. The two girls had mild symptoms and rapid recovery, suggesting that children with SARS-CoV-2 infection may be mild and have a good prognosis. There were differences in the clinical symptoms and imaging findings between the twin girls, suggesting that SARS-CoV-2 infection has diverse clinical features in children.

Supplementation exogenous bile acid improved growth and intestinal immune function associated with NF-κB and TOR signalling pathways in on-growing grass carp (Ctenopharyngodon idella): Enhancement the effect of protein-sparing by dietary lipid.

This study investigated the effects of bile acid (BA) supplementation on growth performance, intestinal immune function and the mRNA expression of the related signalling molecules in on-growing grass carp (Ctenopharyngodon idella). A total of 540 healthy grass carp (mean weight 179.85 ±â€¯1.34 g) were fed a normal protein and lipid (NPNL) diet containing 29% crude protein (CP) and 5% ether extract (EE), and five low-protein and high-lipid (LPHL) diets (26% CP, 6% EE) with graded levels of BA (0-320 mg/kg diet) for 50 days. The fish were then challenged with Aeromonas hydrophila for 14 days. The results indicated that compared with the NPNL diet, the LPHL diet (unsupplemented BA) suppressed the growth performance, intestinal development and enteritis resistance capability and impaired the partial intestinal immune function of on-growing grass carp. Whereas in the LPHL diet, optimal BA supplementation significantly improved fish growth performance (percent weight gain, specific growth rate, feed intake and feed efficiency) and intestinal growth and function (intestine weight, intestine length and intestosomatic index), increased beneficial bacteria Lactobacillus and Bifidobacterium amounts, decreased harmful bacteria Aeromonas and Escherichia coli amounts, elevated lysozyme and acid phosphatase activities, increased complement (C3 and C4) and immunoglobulin M contents, and upregulated ß-defensin-1, hepcidin, liver expressed antimicrobial peptide 2A (LEAP-2A), LEAP-2B, Mucin2, interleukin 10 (IL-10), IL-11, transforming growth factor (TGF)-ß1, TGF-ß2, IL-4/13A (not IL-4/13B), TOR, S6K1 and inhibitor of κBα (IκBα) mRNA levels. In addition, optimal BA supplementation in the LPHL diet downregulated tumour necrosis factor α (TNF-α), interferon γ2 (IFN-γ2), IL-1ß, IL-6, IL-8, IL-15, IL-17D, IL-12p35, IL-12p40 (rather than proximal intestine (PI) or mid intestine (MI), nuclear factor kappa B p65 (NF-κB p65) (except NF-κB p52), c-Rel, IκB kinase ß (IKKß), IKKγ (except IKKα), eIF4E-binding proteins (4E-BP)1 and 4E-BP2 mRNA levels in all three intestinal segments of on-growing grass carp (P < 0.05). These findings suggest that BA supplementation in the LPHL diet improves growth and intestinal immune function of fish. Furthermore, 240 mg/kg BA supplementation in the LPHL diet was superior to the NPNL diet in improving growth and enhancing intestinal immune function of fish. Finally, based on percent weight gain, feed intake, protecting fish against enteritis, lysozyme activity in MI and acid phosphatase activity in distal intestine (DI), the optimal BA supplementation for on-growing grass carp were estimated to be 168.98, 170.23, 166.67, 176.50 and 191.97 mg/kg diet, respectively.

Epigenetics and bone diseases.

Owing to the development of new technologies, the epigenome, a second dimensional method for genome analysis has emerged. Epigenetic mechanisms, including DNA methylation, histone modifications and noncoding RNAs, regulate gene expression without changing the genetic sequence. These epigenetic mechanisms normally modulate gene expression, trans-generational effects and inherited expression states in various biological processes. Abnormal epigenetic patterns typically cause pathological conditions, including cancers, age-related diseases, and specific cartilage and bone diseases. Facing the rapidly developing epigenetic field, we reviewed epigenetic mechanisms and their involvement with the skeletal system and their role in skeletal development, homeostasis and degeneration. Finally, we discuss the prospects for the future of epigenetics.

Polydatin ameliorates Staphylococcus aureus-induced mastitis in mice via inhibiting TLR2-mediated activation of the p38 MAPK/NF-κB pathway.

Recent studies show that Polydatin (PD) extracted from the roots of Polygonum cuspidatum Sieb, a widely used traditional Chinese remedies, possesses anti-inflammatory activity in several experimental models. In this study, we investigated the anti-inflammatory effects of PD on Staphylococcus aureus-induced mastitis in mice and elucidated the potential mechanisms. In mice with S aureus-induced mastitis, administration of PD (15, 30, 45 mg/kg, ip) or dexamethasone (Dex, 5 mg/kg, ip) significantly suppressed the infiltration of inflammatory cells, ameliorated the mammary structural damage, and inhibited the activity of myeloperoxidase, a biomarker of neutrophils accumulation. Furthermore, PD treatment dose-dependently decreased the levels of TNF-α, IL-1ß, IL-6 and IL-8 in the mammary gland tissues. PD treatment also dose-dependently decreased the expression of TLR2, MyD88, IRAK1, IRAK4 and TRAF6 as well as the phosphorylation of TAK1, MKK3/6, p38 MAPK, IκB-α and NF-κB in the mammary gland tissues. In mouse mammary epithelial cells (mMECs) infected by S aureus in vitro, pretreatment with PD dose-dependently suppressed the upregulated pro-inflammatory cytokines and signaling proteins, and the nuclear translocation of NF-κB p65 and AP-1. A TLR2-neutralizing antibody mimicked PD in its suppression on S aureus-induced upregulation of MyD88, p-p38 and p-p65 levels in mMECs. PD (50, 100 µg/mL) affected neither the growth of S aureus in vitro, nor the viability of mMECs. In conclusion, PD does not exhibit antibacterial activity against S aureus, its therapeutic effects in mouse S aureus-induced mastitis depend on its ability to down-regulate pro-inflammatory cytokine levels via inhibiting TLR2-mediated activation of the p38 MAPK/NF-κB signaling pathway.

[Investigation and genetic identification on Babesia infection in rodents in some areas of Fujian Province].

Objective: To explore the status of Babesia infection in rodents and the genetic characteristics of Babesia spp. in Fujian Province. Methods: Rodents were captured by the night trapping method in Shaowu, Qingliu, Shunchang, Yong'an, Changle and Youxi during 2014-2015. The rodent species was identified, and information on the time and place of capture, species and sex of rodents was recorded. Blood samples was collected, in which the fragment of 18S rRNA gene of Babesia spp. was amplified by PCR. The PCR products were sequenced and the phylogenetic tree was constructed for homology analysis. Data on positive rate were analyzed with Chi-square or Fisher exact test. Results: Two hundred and nine rats were captured, comprising of 71 domestic and 138 wild rats. The overall positive rate was 9.6%(20/209). The positive rate in domestic rats was 2.8%(2/71), including one Rattus norvebicus and one Rattus flavipectus. The positive rate in wild rats was 13.0%(18/138), including 13 Bandicota indica, one Rattus losea, 2 Rattus confucianus and 2 Rattus fulvescens. The positive rate was significantly higher in wild rats than in domestic rats (P < 0.05). The Youxi region had the highest positive rate(14.9%,13/87), followed by Yong'an(13.6%, 3/22), and no positive rat was found in Qingliu. The positive rate in the male rats was 7.9%(9/114), and that in the females was 11.6%(11/95). The positive rate was highest in adult rats (10.4%,18/173), followed by young ones (6.3%,2/32). No positive rat was found in old rats. There was no significant difference in positive rate among different regions, between male and female rats, or among different ages (P > 0.05). The sequences of PCR products had a 100% homology. The BLAST results revealed the species to be Babesia microti. The phylogenetic tree showed that the sample sequence was the most homologous with Babesia microti from Zhejiang Province(GenBank Accession No: JQ609305). Conclusion: There occurs Babesia microti infection in rats in part areas of Fujian Province. The positive rate was higher in wild rats than in domestic rats.

Ethnic Differences in Preterm Birth Risks for Pregnant Women with Thyroid Dysfunction or Autoimmunity: A Meta-analysis.

OBJECTIVE: Abnormal maternal thyroid function is associated with preterm birth. However, this association stays dubious in relevant individual studies for ethnic difference reasons and lack of direct supporting data. This study aimed to evaluate the relationship between preterm birth and thyroid dysfunction or autoimmunity based on ethnic differences. METHODS: Relevant studies were identified through searches of MEDLINE, Excerpta Medica, Wan Fang, China Biological Medicine disc, and China National Knowledge Infrastructure from inception to June 15, 2016. Original articles in which an incidence or prevalence of thyroid dysfunction or autoimmunity before second trimester of pregnancy could be extracted were included. RESULTS: Thirty-two unique studies were included for the final meta-analysis. Patients involved were divided into two groups: Group 1 (G1) and Group 2 (G2) comprising of Asian and Caucasian populations, respectively. Positive thyroid antibodies were associated with the occurrence of preterm birth in both G1 [odds ratio (OR): 3.62, 95% confidence interval (CI): 2.83-4.65] and G2 (OR: 1.35, 95% CI: 1.17-1.56); hypothyroidism, only in G2 (OR: 1.20, CI: 1.09-1.33); and subclinical hypothyroidism or hypothyroxinemia, in neither group. CONCLUSION: Thyroid autoimmunity may be a more favorable factor leading to preterm birth among pregnant women of different ethnicities, compared with thyroid dysfunction.

[Application of time domain reflectometry for determination of wate content in Xiangsha Yangwei pills].

Xiangsha Yangwei pill was selected as a model drug in this research, and time domain reflectometry (TDR) was used to determine the water content in the pill. The effects of five factors including the number of pill layers, pill packing density, atmospheric moisture, ambient temperature and the ratio of pill formula were investigated on water content. The results showed that the number of pill layers and ambient temperature had significant effects on water content of pills, while the pill packing density, atmospheric moisture and pill formula ratio had little effect on the determination of water content in pills. The reflection value was stable when 6 layers of pills were used. Under the condition of 25 ℃ and 45% relative humidity, the water content of pills ranged from 4.01% to 22.38%, showing good linear relationship between water content and reflection value, and the model equation was as follows: Y=0.279X－21.670 (R²=0.997 0). Verification experiment was used to explain the feasibility of this prediction model. The precision of the method complied with the methodology standard. It is concluded that TDR can be used in determination of water content in Xiangsha Yangwei pills. Additionally, TDR, as a new way to quickly and efficiently determine the water content, has a prospect application in the processing of traditional Chinese medicine pharmacy, especially for concentrated pill.

Interaction of gas phase oxalic acid with ammonia and its atmospheric implications.

Oxalic acid is believed to play an important role in the formation and growth of atmospheric organic aerosols. However, as a common organic acid, the understanding of the larger clusters formed by gas phase oxalic acid with multiple ammonia molecules is incomplete. In this work, the structural characteristics and thermodynamics of oxalic acid clusters with up to six ammonia molecules have been investigated at the PW91PW91/6-311++G(3df,3pd) level of theory. We found that oxalic acid forms relatively stable clusters with ammonia molecules, and that ionization events play a key role. The analyses of the thermodynamics and atmospheric relevance indicate that the heterodimer (H2C2O4)(NH3) shows an obvious relative concentration in the atmosphere, and thus likely participates in new particle formation. However, with increasing number of ammonia molecules, the concentration of clusters decreases gradually. Additionally, clusters of oxalic acid with ammonia molecules are predicted to form favorably in low temperature conditions and show high Rayleigh scattering intensities.

A Case-control Study of Environmental Risk Factors for Nonsyndromic Cleft of the Lip and/or Palate in Xuzhou, China.

In this study, we sought to determine the association between environmental factors and nonsyndromic cleft of the lip and/or palate (NSCLP) to understand the etiology of the disease. A total of 200 NSCLP cases and 327 controls were recruited at the Maternal and Child Health Hospital of Xuzhou City. We conducted face-to-face interviews with the mothers of both cases and controls. The factors increasing the risk of NSCLP were a positive family history [odds ratio (OR)=56.74], pesticide exposure (OR=8.90), and indoor decoration pollution (OR=4.32). On the other hand, the factors decreasing the risk of NSCLP were a high education level (OR=0.22) and supplementation of folic acid (OR=0.23) and multivitamins (OR=0.16). Positive family history, pesticide exposure, and indoor decoration pollution are associated with the risk of NSCLP. In contrast, high education level and folic acid and multivitamin supplementation are protective factors against NSCLP.

Identification of susceptibility genes in non-syndromic cleft lip with or without cleft palate using whole-exome sequencing.

BACKGROUND: Non-syndromic cleft lip with or without cleft palate (NSCL/P) is among the most common congenital malformations. The etiology of NSCL/P remains poorly characterized owing to its complex genetic heterogeneity. The objective of this study was to identify genetic variants that increase susceptibility to NSCL/P. MATERIAL AND METHODS: Whole-exome sequencing (WES) was performed in 8 fetuses with NSCL/P in China. Bioinformatics analysis was performed using commercially available software. Variants detected by WES were validated by Sanger sequencing. RESULTS: By filtering out synonymous variants in exons, we identified average 8575 nonsynonymous single nucleotide variants (SNVs). We subsequently compared the SNVs against public databases including NCBI dbSNP build 135 and 1000 Genomes Project and obtained an average of 203 SNVs. Total 12 reported candidate genes were verified by Sanger sequencing. Sanger sequencing also confirmed 16 novel SNVs shared by two or more samples. CONCLUSIONS: We have found and confirmed 16 susceptibility genes responsible for NSCL/P, which may play important role in the etiology of NSCL/P. The susceptibility genes identified in this study will not only be useful in revealing the etiology of NSCL/P but also in diagnosis and treatment of the patients with NSCL/P.

TLR4-dependant immune response, but not hepatitis B virus reactivation, is important in radiation-induced liver disease of liver cancer radiotherapy.

Toll-like receptor 4 (TLR4) is an important trigger of the immune response against hepatitis B virus (HBV) infection and liver injuries. The roles of HBV reactivation versus TLR4-dependant immune response may be critical factors in preventing radiation-induced liver diseases (RILDs) after liver cancer radiotherapy. This study consists of three phases. In the primary phase, livers of mutant TLR4 (TLR4(-)) mice were irradiated with 30 Gy in either the absence or presence of HBV infection. The latter was done by introduction of plasmid pAAV/HBV 1.2. In the advanced phase, RILDs were compared in normal TLR4 (TLR4(+)) versus TLR4(-) mice. In the validation phase, 28 liver cancer patients who had undergone radiotherapy before hepatectomy were enrolled. Liver biopsies near tumors, irradiated with 35-48 Gy, were used to construct tissue microarrays. HBV reactivation, TLR4 expression, and severity of RILDs were studied in both mouse and human. More HBV reactivation, without significant RILD, was observed in irradiated versus unirradiated TLR4(-) mice. RILD scores of TLR4(+) mice were higher than TLR4(-) mice. In humans, serious RILDs tended to develop in patients with high TLR4 expression, but not in patients with low TLR4 or high HBV surface antigen expression. High TLR4 expression was seen in only 2 of 12 HBV-reactive patients, but in HBV-nonreactive patients, it was seen in 6 of 9 (P < 0.03). In summary, RILDs correlated with high TLR4 expression, but not with HBV reactivation, which is inhibited in liver with high TLR4 expression after liver cancer radiotherapy.

[Near infrared spectroscopy synergy interval wavelength selection method using the LSSVM model].

The present paper proposes a wavelength selection algorithm based on nonlinear factors named Synergy interval least squares support vector machines (siLSSVM). siLSSVM combines the interval strategy of wavelength selection method with the idea of synergy interval and overcomes the disadvantages of the traditional wavelength selection methods, i. e. ignoring the nonlinear factors. Taking the near infrared spectrum data of apple sugar as performance verification object of this new algorithm, comparing new algorithm with siPLS, the model performance has been greatly improved. The root-mean-square error (RMSEP) in new algorithm has increased respectively by 37.43% and 47.88% under the model of PLS and LSSVM, with increases of 6.04% and 7.31% in the correlative coefficient (RP). The examples illustrate that siLSSVM can efficiently select the optimum wavelength interval for spectrum data with strong nonlinear factors. This algorithm greatly improves the prediction accuracy and robustness of the model, which provides a new prospect for near infrared spectral with nonlinear factors to select wavelength.

Genetic mutation of Tas2r104/Tas2r105/Tas2r114 cluster leads to a loss of taste perception to denatonium benzoate and cucurbitacin B.

BACKGROUND: Bitter taste receptors (Tas2rs) are generally considered to sense various bitter compounds to escape the intake of toxic substances. Bitter taste receptors have been found to widely express in extraoral tissues and have important physiological functions outside the gustatory system in vivo. METHODS: To investigate the physiological functions of the bitter taste receptor cluster Tas2r106/Tas2r104/Tas2r105/Tas2r114 in lingual and extraoral tissues, multiple Tas2rs mutant mice and Gnat3 were produced using CRISPR/Cas9 gene-editing technique. A mixture containing Cas9 and sgRNA mRNAs for Tas2rs and Gnat3 gene was microinjected into the cytoplasm of the zygotes. Then, T7EN1 assays and sequencing were used to screen genetic mutation at the target sites in founder mice. Quantitative real-time polymerase chain reaction (qRT-PCR) and immunostaining were used to study the expression level of taste signaling cascade and bitter taste receptor in taste buds. Perception to taste substance was also studied using two-bottle preference tests. RESULTS: We successfully produced several Tas2rs and Gnat3 mutant mice using the CRISPR/Cas9 technique. Immunostaining results showed that the expression of GNAT3 and PLCB2 was not altered in Tas2rs mutant mice. But qRT-PCR results revealed the changed expression profile of mTas2rs gene in taste buds of these mutant mice. With two-bottle preference tests, these mutant mice eliminate responses to cycloheximide due to genetic mutation of Tas2r105. In addition, these mutant mice showed a loss of taste perception to quinine dihydrochloride, denatonium benzoate, and cucurbitacin B (CuB). Gnat3-mediated taste receptor and its signal pathway contribute to CuB perception. CONCLUSIONS: These findings implied that these mutant mice would be a valuable means to understand the biological functions of TAS2Rs in extraoral tissues and investigate bitter compound-induced responses mediated by these TAS2Rs in many extraoral tissues.

A Novel Cargo Delivery System-AnCar-ExoLaIMTS Ameliorates Arthritis via Specifically Targeting Pro-Inflammatory Macrophages.

Macrophages are heterogenic phagocytic cells that play distinct roles in physiological and pathological processes. Targeting different types of macrophages has shown potent therapeutic effects in many diseases. Although many approaches are developed to target anti-inflammatory macrophages, there are few researches on targeting pro-inflammatory macrophages, which is partially attributed to their non-s pecificity phagocytosis of extracellular substances. In this study, a novel recombinant protein is constructed that can be anchored on an exosome membrane with the purpose of targeting pro-inflammatory macrophages via antigen recognition, which is named AnCar-ExoLaIMTS . The data indicate that the phagocytosis efficiencies of pro-inflammatory macrophages for different AnCar-ExoLaIMTS show obvious differences. The AnCar-ExoLaIMTS3 has the best targeting ability for pro-inflammatory macrophages in vitro and in vivo. Mechanically, AnCar-ExoLaIMTS3 can specifically recognize the leucine-rich repeat domain of the TLR4 receptor, and then enter into pro-inflammatory macrophages via the TLR4-mediated receptor endocytosis pathway. Moreover, AnCar-ExoLaIMTS3 can efficiently deliver therapeutic cargo to pro-inflammatory macrophages and inhibit the synovial inflammatory response via downregulation of HIF-1α level, thus ameliorating the severity of arthritis in vivo. Collectively, the work established a novel gene/drug delivery system that can specifically target pro-inflammatory macrophages, which may be beneficial for the treatments of arthritis and other inflammatory diseases.

Temporospatial expression of Dmrt1 in chicken urogenital system (Gallus gallus) using whole mount in situ hybridization.

Doublesex and mab-3-related transcription factor 1 (Dmrt1) is a Z-linked gene that putatively determines the phenotype of gonads in birds. The sex differential expression of Dmrt1 was examined using wholemount in situ hybridization (WISH) in the urogenital systems during embryogenesis. The results revealed that Dmrt1 showed dimorphic expression in chicken gonads, which increased from day 6.5 to day 10.5. The expression of Dmrt1 in male (ZZ) gonads was not twice as much as in female (ZW) gonads, suggesting the existence of other regulatory mechanisms in addition to Z chromosome dosage effect.

[Corneal collagen cross-linking in the treatment of progressive keratoconus-preliminary results].

OBJECTIVE: To evaluate the efficacy and safety of riboflavin-ultraviolet-A (UV-A)-induced corneal collagen cross-linking (CXL) in the management of progressive keratoconus. METHODS: It was a retrospective case series study. Twenty-three eyes of 13 patients with progressive keratoconus were included. Corneal collagen crosslinking was performed under topical anesthesia including corneal de-epithelization (8 mm diameter) and instillation of 0.1% riboflavin (in 20% dextran T500 solution) every 3 minutes for a total of 30 minutes. The irradiation is performed for another 30 min using a solid-state UV-A illuminator at 370 nm and an irradiance of 3 mW/cm(2). Average follow-up was 15.23 ± 3.39 months (range: 12 to 22 months). Visual acuity, corneal topography, in vivo confocal microscopy, and endothelial cell count were evaluated at baseline and at 1, 3, 6, and 12 months follow-up. RESULTS: Mean uncorrected visual acuity(UCVA) and best spectacle-corrected visual acuity (BSCVA) increased 0.115 ± 0.158 LogMAR (t = 3.418, P = 0.0026) and 0.114 ± 0.218 LogMAR (t = 2.441, P = 0.0236) 12 months postoperatively respectively. Interim analysis of treated eyes showed a flattening of the steepest simulated keratometry value (K-max) and astigmatism by an average of (1.893 ± 3.713) diopters (D) (t = 2.391, P = 0.0262) and (0.117 ± 1.488) D (t = 0.370, P = 0.715) respectively at 12 months. Central corneal thickness decreased by (27.5 ± 26.8) µm (t = 4.812, P = 0.000) and (1.54 ± 19.4) µm ( t = 0.147, P = 0.885) at one month and 12 months postoperatively respectively.Intraocular pressure, endothelial cell count, lens and fundus didn't change significantly at 12 months follow-up. CONCLUSIONS: CXL stabilised and improved the UCVA and BSCVA as well as the maximum k-readings at 1 year postoperatively in our cohort. It seems to be a safe and promising procedure to stop the progression of keratoconus.

CircPRRC2C Promotes the Oncogenic Phenotypes of Laryngeal Squamous Cell Carcinoma Cells via MiR-136-5p/HOXD11 Axis.

The involvement of circular RNAs (circRNAs) in laryngeal squamous cell carcinoma (LSCC) carcinogenesis has gradually been proposed. Herein, we aimed to explore the function and mechanism of circPRRC2C in LSCC. Quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting were used for detecting the content of genes and proteins. In vitro experiments were conducted using 5-ethynyl-2'-deoxyuridine, colony formation, flow cytometry, and transwell assays. The binding between miR-136-5p and circPRRC2C or Homeobox D11 (HOXD11) was confirmed by using the dual-luciferase reporter assay. The murine xenograft model was established for in vivo analysis. The commercial kit was used for exosome separation. CircPRRC2C is a stable circRNA, and was highly expressed in LSCC tissues and cell lines. Functionally, circPRRC2C deficiency impaired LSCC cell proliferation, migration and invasion but induced cell apoptosis in vitro and impeded tumor growth in vivo, however, circPRRC2C overexpression showed the exact opposite effects. Mechanistically, circPRRC2C directly targeted miR-136-5p, which showed inhibitory effects on the growth and mobility of LSCC cells. Meanwhile, miR-136-5p directly targeted HOXD11, and circPRRC2C/miR-136-5p/HOXD11 formed a feedback loop in LSCC cells. Further rescue assays exhibited that circPRRC2C exerted its effects by miR-136-5p/HOXD11 axis. In addition, circPRRC2C was stably packaged into exosomes and showed potential diagnostic value for LSCC. CircPRRC2C acted as an oncogene to promote LSCC cell oncogenic phenotypes via miR-136-5p/HOXD11 axis, besides, circPRRC2C was stably packaged into exosomes, indicating the potential application of circPRRC2C-targeting agents in the treatment in LSCC.