[In vitro expression and functional analyses of the mutants p.R243Q, p.R241C and p.Y356X of the human phenylalanine hydroxylase]. / äººè‹¯ä¸™æ°¨é ¸ç¾ŸåŒ–é ¶çªå˜ä½“p.R243Q、p.R241C和p.Y356Xçš„ä½“å¤–è¡¨è¾¾åŠåŠŸèƒ½ç ”ç©¶.

OBJECTIVES: To study the in vitro expression of three phenylalanine hydroxylase (PAH) mutants (p.R243Q, p.R241C, and p.Y356X) and determine their pathogenicity. METHODS: Bioinformatics techniques were used to predict the impact of PAH mutants on the structure and function of PAH protein. Corresponding mutant plasmids of PAH were constructed and expressed in HEK293T cells. Quantitative reverse transcription polymerase chain reaction was used to measure the mRNA expression levels of the three PAH mutants, and their protein levels were assessed using Western blot and enzyme-linked immunosorbent assay. RESULTS: Bioinformatics analysis predicted that all three mutants were pathogenic. The mRNA expression levels of the p.R243Q and p.R241C mutants in HEK293T cells were similar to the mRNA expression level of the wild-type control (P>0.05), while the mRNA expression level of the p.Y356X mutant significantly decreased (P<0.05). The PAH protein expression levels of all three mutants were significantly reduced compared to the wild-type control (P<0.05). The extracellular concentration of PAH protein was reduced in the p.R241C and p.Y356X mutants compared to the wild-type control (P<0.05), while there was no significant difference between the p.R243Q mutant and the wild type control (P>0.05). CONCLUSIONS: p.R243Q, p.R241C and p.Y356X mutants lead to reduced expression levels of PAH protein in eukaryotic cells, with p.R241C and p.Y356X mutants also affecting the function of PAH protein. These three PAH mutants are to be pathogenic.

Assisted reproductive technology treatment failure and the detection of intrauterine HPV through spent embryo transfer media sample.

Cervical human papillomavirus (HPV) infection is believed to increase the risks of pregnancy failure and abortion, however, whether the uterine cavity HPV infection reduces pregnancy rate or increases miscarriage rate remains unclarified in infertile women undergoing assisted reproductive technology (ART) treatment. Therefore, we aimed to assess ART outcomes in the presence of intrauterine HPV. This was a hospital-based multicenter (five reproductive medicine centers) matched cohort study. This study involved 4153 infertile women undergoing in vitro fertilization (IVF) or intracytoplasmic sperm injection treatment in five reproductive medicine centers between October 2018 and 2020. The spent embryo transfer media sample with endometrium tissue were collected and performed with flow-through hybridization and gene chips to detect HPV DNA. According to basic characteristics, HPV-positive and negative patients were matched in a ratio of 1:4 by age, body mass index transfer timing, transfer type, and number of embryos transferred. The primary outcome was pregnancy and clinical miscarriage rates in the transfer cycle underwent HPV detection. 92 HPV-positive and 368 HPV-negative patients were screened and analyzed statistically. Univariate analysis showed uterine cavity HPV infection resulted in lower rates of ongoing pregnancy (31.5% vs. 44.6%; p = 0.023), implantation (32.3% vs. 43.1%; p = 0.026), biochemical pregnancy (47.8% vs. 62.5%; p = 0.010), and clinical pregnancy (40.2% vs. 54.3%; p = 0.015) compared with HPV negative group. The infertile female with positive HPV also had a slightly higher frequency of biochemical miscarriage (15.9% vs. 13.0%; p = 0.610) and clinical miscarriage (24.3% vs. 15.5%; p = 0.188). These findings suggest that HPV infection in the uterine cavity is a high risk for ART failure. HPV screening is recommended before ART treatment, which may be benefit to improving pregnancy outcome.

Development of an artificial intelligence based model for predicting the euploidy of blastocysts in PGT-A treatments.

The euploidy of embryos is unpredictable before transfer in in vitro fertilisation (IVF) treatments without pre-implantation genetic testing (PGT). Previous studies have suggested that morphokinetic characteristics using an artificial intelligence (AI)-based model in the time-lapse monitoring (TLM) system were correlated with the outcomes of frozen embryo transfer (FET), but the predictive effectiveness of the model for euploidy remains to be perfected. In this study, we combined morphokinetic characteristics, morphological characteristics of blastocysts, and clinical parameters of patients to build a model to predict the euploidy of blastocysts and live births in PGT for aneuploidy treatments. The model was effective in predicting euploidy (AUC = 0.879) but was ineffective in predicting live birth after FET. These results provide a potential method for the selection of embryos for IVF treatments with non-PGT.

Feeding intolerance alters the gut microbiota of preterm infants.

Feeding intolerance (FI) is a common disease in preterm infants, often causing a delay in individual development. Gut microbiota play an important role in nutrient absorption and metabolism of preterm infants. To date, few studies have focused on the community composition of gut microbiota of preterm infants with feeding intolerance. In this study, we collected fecal samples from 41 preterm infants diagnosed with feeding intolerance and 29 preterm infants without feeding intolerance, at three specific times during the development and prevalence of feeding intolerance (after birth, when feeding intolerance was diagnosed, after feeding intolerance was gone), from different hospitals for 16S rRNA gene sequencing. The gut microbiota community composition of preterm infants diagnosed with feeding intolerance was significantly different from that of preterm infants without feeding intolerance. At the time when feeding intolerance was diagnosed, the relative abundance of Klebsiella in preterm infants with feeding intolerance increased significantly, and was significantly higher than that of the preterm infants without feeding intolerance. After feeding intolerance was cured, the relative abundance of Klebsiella significantly decreased in the infants diagnosed with feeding intolerance, while the relative abundance of Klebsiella in preterm infants without feeding intolerance was not significantly altered during the development and prevalence of feeding intolerance. Furthermore, we verified that Klebsiella was effective in the diagnosis of feeding intolerance (AUC = 1) in preterm infants, suggesting that Klebsiella is a potential diagnostic biomarker for feeding intolerance.

Comparative Analysis of Gut Microbial Communities in Children under 5 Years Old with Diarrhea.

Diarrhea is a global disease with a high morbidity and mortality rate in children. In this study, 25 fecal samples were collected from children under 5 years old. Seven samples had been taken from healthy children without diarrhea and marked as the healthy control group; eight samples had been sampled from children with diarrhea caused by dyspepsia and defined as the non-infectious group; and ten samples had been taken from children with diarrhea induced by intestinal infections and identified as the infectious group. We detected the microbial communities of samples by using high-throughput sequencing of 16S rRNA genes. The proportion of aerobic and facultative anaerobic microbes in samples of the infectious group was much higher than in the non-infectious group. In addition, the relative abundance of Enterococcus in the healthy control group was significantly higher than in the non-infectious group and infectious group. This can be used as a potential diagnostic biomarker for diarrhea.

Immobilization of collagen peptide on dialdehyde bacterial cellulose nanofibers via covalent bonds for tissue engineering and regeneration.

Bacterial cellulose (BC) is an alternative nanostructured biomaterial to be utilized for a wide range of biomedical applications. Because of its low bioactivity, which restricted its practical application, collagen and collagen hydrolysate were usually composited into BC. It is necessary to develop a new method to generate covalent bonds between collagen and cellulose to improve the immobilization of collagen on BC. This study describes a facile dialdehyde BC/collagen peptide nanocomposite. BC was oxidized into dialdehyde bacterial cellulose (DBC) by regioselective oxidation, and then composited with collagen peptide (Col-p) via covalent bonds to form Schiff's base type compounds, which was demonstrated by the results of microstructures, contact angle, Col-p content, and peptide-binding ratio. The peptide-binding ratio was further affected by the degree of oxidation, pH value, and zeta potential. In vitro desorption measurement of Col-p suggested a controlled release mechanism of the nanocomposite. Cell tests indicated that the prepared DBC/Col-p composite was bioactive and suitable for cell adhesion and attachment. This work demonstrates that the DBC/Col-p composite is a promising material for tissue engineering and regeneration.