The screening and expression of polysaccharide deacetylase from Caulobacter crescentus and its function analysis.

The bacterium Caulobacter crescentus secretes an adhesive polysaccharide called holdfast, which is the known strongest underwater adhesive in nature. The deacetylase encoded by hfs (holdfast synthesis) H gene is a key factor affecting the adhesion of holdfast. Its structure and function are not yet clear, and whether other polysaccharide deacetylases exist in C. crescentus is still unknown. The screening of both HfsH and its structural analogue as well as their purification from the artificial expression products of Escherichia coli is the first step to clarify these questions. Here, we determined the conserved domains of HfsH via sequence alignment among carbohydrate esterase family 4 enzymes and screened out its structural analogue (CC_2574) in C. crescentus. The recombinant HfsH and CC_2574 were effectively expressed in E. coli. Both of them were purified by chromatography from their corresponding productions in E. coli and were then functionally analyzed. The results indicated that a high deacetylase activity (61.8 U/mg) was observed in recombinant HfsH but not in CC_2574, which suggesting that HfsH might be the irreplaceable gene mediating adhesion of holdfast in C. crescentus. Moreover, the divalent metal ions Zn2+ , Mg2+ , and Mn2+ could promote the activity of recombinant HfsH at the concentration from 0.05 to 1 mM, but inhibit its activity when the concentration exceeds 1 mM. In sum, our study first realized the artificial production of polysaccharide deacetylase HfsH and its structural analogue, and further explored their functions, both of which laid the foundation for the development of new adhesive materials.

Surgical management and outcome of primary intracranial Rosai-Dorfman disease: a single-institute experience and pooled analysis of individual patient data.

Primary intracranial Rosai-Dorfman disease (PIRDD) is considered a nonmalignant nonneoplastic entity, and the outcome is unclear due to its rarity. The study aimed to elaborate the clinic-radiological features, treatment strategies, and progression-free survival (PFS) in patients with PIRDD. Patients with pathologically confirmed PIRDD in our institute were reviewed. Literature of PIRDD, updated until December 2019, was systematically searched in 7 databases (Embase, PubMed, Cochrane database, Web of Science, Wanfang Data Knowledge Service Platform, the VIP Chinese Science and Technology Periodical Database (VIP), and the China National Knowledge Infrastructure (CNKI)). These prior publication data were processed and used according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. Clinical-radiological characteristics and adverse factors for PFS were evaluated in the pooled cohort. The pooled cohort of 124 cases (81 male and 43 female), with a mean age of 39.7 years, included 11 cases from our cohort and 113 cases from 80 prior studies. Twenty-nine patients (23.4%) had multiple lesions. Seventy-four patients (59.7%) experienced gross total resection (GTR), 50 patients (40.3%) had non-GTR, 15 patients (12.1%) received postoperative adjuvant radiation, and 23 patients (18.5%) received postoperative steroids. A multivariate Cox regression revealed that GTR (HR = 4.52; 95% CI 1.21-16.86; p = 0.025) significantly improved PFS, and multiple lesions (p = 0.060) tended to increase the hazard of recurrence. Neither radiation (p = 0.258) nor steroids (p = 0.386) were associated with PFS. The overall PFS at 3, 5, and 10 years in the pooled cohort was 88.4%, 79.4%, and 70.6%, respectively. The PFS at 5 and 10 years in patients with GTR was 85.4% and 85.4%, respectively, which was 71.5% and 35.8%, respectively, in patients without GTR. Gross total resection significantly improved PFS and was recommended for PIRDD. Radiation and steroids were sometimes empirically administered for residual, multiple, or recurrent PIRDD, but the effectiveness remained arguable and required further investigation.Systematic review registration number: CRD42020151294.

PI3K-AKT-mTOR signaling pathway: the intersection of allergic asthma and cataract.

Allergic asthma is a chronic inflammatory disease and involves many cells and cellular components. Cataract is a condition that affects the transparency of the lens, which the opacity of the lens caused by any innate or acquired factor degrades its transparency or changes in color. During the establishment of asthma model of rats with chicken ovalbumin nebulization, it was found that asthmatic rats were more likely to have monocular or binocular cataract symptoms than normal rats. Considering that they are all induced by immune imbalance, inflammation, etc., there may be some correlation in the mechanism, and many clues showed that both diseases are associated with activation of the PI3K-AKT-mTOR signaling pathway. Therefore, we hypothesized that the PI3K-AKT-mTOR signaling pathway produces inflammatory or immune imbalance based on allergy leading to cataract.

Construction and identification of the recombinant hFcεRIα/RBL-2H3 cells.

IgE/FcεRI signal pathway plays a crucial role in triggering allergic reactions, and there is no cross-recognition between IgE and FcεRI in human and rats. In order to obtain the hFcεRIα/ RBL-2H3 cell line, total RNA was extracted from U937 cells, and the human FcεRIα gene was obtained by RT-PCR technology. Then the amplified product was digested and inserted into the pIRES2-EGFP vector. After the plasmid was transfected into the RBL-2H3 cells using lipofectamine, and the RBL-2H3 cell lines of stable expression were screened by G418. The transfection efficiency reached 60.45% with optimizing transfection parameters. The last the expression of hFcεRIα was detected by RT-PCR, western blotting and fluorescent microscopy. The present results demonstrated that the pIRES2-EGFP-hFcεRIα vector was constructed and a stable cell line of hFcεRIα/ RBL-2H3 cells was established successfully. This cell line is promising tools for further research on the pathogenesis and drug development of allergic diseases.

Combining with volatilomic profiling and chemometrics to explore the volatile characteristics in five different dried Zanthoxylum bungeanum maxim.

Owing to the short picking period of the fresh Zanthoxylum bungeanum, the postharvest drying has become an essential operation before the storage and transportation of Z. bungeanum. To explore the effects of drying methods on volatile characteristics, the volatilomic profiling of five different dried Z. bungeanum was investigated by E-nose, HS-SPME-GC/MS, GC-IMS in combination with chemometrics. The results indicated that W1W, W2W and W5S sensors within E-nose analysis showed the strongest responses in both fresh and dried Z. bungeanum. According to the identification of volatile organic compounds (VOCs), terpenes, esters and alcohols played the major roles in the volatile formation of the fresh and dried Z. bungeanum. The samples derived from hot air drying showed the relatively similar features with the fresh sample based on the relative abundances of these major VOCs. According to the results of multiple factor analysis (MFA), GC-IMS showed the strongest ability in distinguishing the fresh and different dried samples. Compared with the high levels of terpenes in fresh group, the significant increasement of terpene alcohols and terpene esters from the degradation and transformation of bound terpenoids was the main characteristics of all dried Z. bungeanum. Using the GC-IMS datasets, a weighted correlation network analysis (WCNA) model was constructed to clarify the VOC characteristics in all detetected samples. Thereinto, 6 significantly correlated modules were identified in fresh and five different dried samples. Additionally, a total of 23 hub VOCs can be recognized as the potential biomarkers for better distinguishing the fresh and five different dried Z. bungeanum.

Determinants of acute and subacute case-fatality in elderly patients with hypertensive intracerebral hemorrhage.

Background: Given that limited reports have described the survival and risk factors for elderly patients with hypertensive intracerebral hemorrhage (HICH), we aimed to develop a valid but simple prediction nomogram for the survival of HICH patients. Methods: All elderly patients ≥65 years old who were diagnosed with HICH between January 2011 and December 2019 were identified. We performed the least absolute shrinkage and selection operator (Lasso) on the Cox regression model with the R package glmnet. A concordance index was performed to calculate the nomogram discrimination; and calibration curves and decision curves were graphically evaluated by depicting the observed rates against the probabilities predicted by the nomogram. Results: A total of 204 eligible patients were analyzed, and over 20 % of the population was above the age of 80 (65-79 years old, n = 161; 80+ years old, n = 43). A hematoma volume ≥13.64 cm3 was associated with higher 7-day mortality (OR = 6.773, 95 % CI = 2.622-19.481; p < 0.001) and higher 90-day mortality (OR = 3.955, 95 % CI = 1.611-10.090, p = 0.003). A GCS score between 13 and 15 at admission was associated with a 7-day favorable outcome (OR = 0.025, 95 % CI = 0.005-0.086; p < 0.001) and a 90-day favorable outcome (OR = 0.033, 95 % CI = 0.010-0.099; p < 0.001). Conclusions: Our nomogram models were visualized and accurate. Neurosurgeons could use them to assess the prognostic factors and provide advice to patients and their relatives.

3D-printed collagen/silk fibroin/secretome derived from bFGF-pretreated HUCMSCs scaffolds enhanced therapeutic ability in canines traumatic brain injury model.

The regeneration of brain tissue poses a great challenge because of the limited self-regenerative capabilities of neurons after traumatic brain injury (TBI). For this purpose, 3D-printed collagen/silk fibroin/secretome derived from human umbilical cord blood mesenchymal stem cells (HUCMSCs) pretreated with bFGF scaffolds (3D-CS-bFGF-ST) at a low temperature were prepared in this study. From an in vitro perspective, 3D-CS-bFGF-ST showed good biodegradation, appropriate mechanical properties, and good biocompatibility. In regard to vivo, during the tissue remodelling processes of TBI, the regeneration of brain tissues was obviously faster in the 3D-CS-bFGF-ST group than in the other two groups (3D-printed collagen/silk fibroin/secretome derived from human umbilical cord blood mesenchymal stem cells (3D-CS-ST) group and TBI group) by motor assay, histological analysis, and immunofluorescence assay. Satisfactory regeneration was achieved in the two 3D-printed scaffold-based groups at 6 months postsurgery, while the 3D-CS-bFGF-ST group showed a better outcome than the 3D-CS-ST group.

[Effect of cotton-moxibustion on chronic eczema and quality of life].

OBJECTIVE: To compare the therapeutic effect between cotton-moxibustion and compound flumetasone ointment, and observe the effect on quality of life in patients with chronic eczema. METHODS: A total of 66 patients with chronic eczema were randomized into an observation group (33 cases, 2 cases dropped off) and a control group (33 cases, 2 cases dropped off). In the observation group, cotton-moxibustion was adopted on target skin lesion, once a day, 3 cones a time. In the control group, external application of compound flumetasone ointment was given twice a day. The treatment for 3 weeks was required in the both groups. Before treatment and 1,2,3 weeks into treatment, scores of visual analogue scale (VAS), eczema area and severity index (EASI) and dermatology life quality index (DLQI) were observed, and the recurrence rate was evaluated in the follow-up one month after treatment. RESULTS: Compared before treatment, the VAS scores of 1,2,3 weeks into treatment, the EASI and DLQI scores of 2,3 weeks into treatment were decreased in the both groups (P<0.05), and those in the observation group were lower than the control group (P<0.05). The follow-up recurrence rate in the observation group were lower than the control group (P<0.05). CONCLUSION: Cotton-moxibustion can effectively improve the pruritus symptom, skin lesion and quality of life in the patients with chronic eczema, the therapeutic effect is superior to the external application of compound flumetasone ointment.

Molecular analysis of phosphomannomutase (PMM) genes reveals a unique PMM duplication event in diverse Triticeae species and the main PMM isozymes in bread wheat tissues.

BACKGROUND: Phosphomannomutase (PMM) is an essential enzyme in eukaryotes. However, little is known about PMM gene and function in crop plants. Here, we report molecular evolutionary and biochemical analysis of PMM genes in bread wheat and related Triticeae species. RESULTS: Two sets of homologous PMM genes (TaPMM-1 and 2) were found in bread wheat, and two corresponding PMM genes were identified in the diploid progenitors of bread wheat and many other diploid Triticeae species. The duplication event yielding PMM-1 and 2 occurred before the radiation of diploid Triticeae genomes. The PMM gene family in wheat and relatives may evolve largely under purifying selection. Among the six TaPMM genes, the transcript levels of PMM-1 members were comparatively high and their recombinant proteins were all enzymatically active. However, PMM-2 homologs exhibited lower transcript levels, two of which were also inactive. TaPMM-A1, B1 and D1 were probably the main active isozymes in bread wheat tissues. The three isozymes differed from their counterparts in barley and Brachypodium distachyon in being more tolerant to elevated test temperatures. CONCLUSION: Our work identified the genes encoding PMM isozymes in bread wheat and relatives, uncovered a unique PMM duplication event in diverse Triticeae species, and revealed the main active PMM isozymes in bread wheat tissues. The knowledge obtained here improves the understanding of PMM evolution in eukaryotic organisms, and may facilitate further investigations of PMM function in the temperature adaptability of bread wheat.

Integrative Exome Sequencing Analysis in Castration-Resistant Prostate Cancer in Chinese Population.

BACKGROUND: Castration-resistant Prostate Cancer (CRPC) is a fatal disease with rapid growth. The malignancy usually presents with metastasis and poor prognosis, and causes 100% mortality. Therefore, the treatment of CRPC is extremely challenging, and its pathogenesis need to be elucidated urgently. OBJECTIVE: The high throughput sequencing technology was used to sequence the whole exome associated with CRPC, to explore the molecular mechanism of CRPC, and to find the potential therapeutic targets. METHODS: We performed whole-exome sequencing of FFPE tissue from 11 Chinese adult male patients. Genomic DNA was fragmented and enriched for whole-exome sequencing using the QiAamp DNA FFPE Tissue KIT, sequenced on an Illumina HiSeq2000 platform, and the relevant genes were analyzed using biological information. Finally, immunohistochemistry method was used to detect the phosphorylation level of LATS1 in CRPC tissues of MST1 mutant and non-mutant patients. RESULTS: We have screened 85 significant mutant genes with relatively high mutation rates of TP53, AR, KMT2, DMAPK1, PIK3R1, SH2B3, WHSC1, KMT2D, MST1 and MAPK1. We first found that MST1 has multiple mutations in CRPC patients, and the MST1 plays an important role in the Hippo pathway. Immunohistochemistry results showed that the phosphorylation level of LATS1 in the mutant patients was significantly lower than that in the non-mutant patients. CONCLUSION: We speculate that MST1 would be a new potential target for the treatment of CRPC by regulating Hippo signaling pathway. The results provided an important clue to the molecular mechanism of CRPC.

Cross-talk of Signaling Pathways in the Pathogenesis of Allergic Asthma and Cataract.

Allergic asthma is a chronic inflammatory disease, which involves many cellular and cellular components. Cataract is a condition that affects the transparency of the lens, which the opacity of the lens caused by any innate or acquired factor degrades its transparency or changes in color. Both of them belong to diseases induced by immune disorders or inflammation. We want to confirm the signaling pathways involved in the regulation of asthma and cataract simultaneously, and provide reference for the later related experiments. So we conducted a scoping review of many databases and searched for studies (Academic research published in Wiley, Springer and Bentham from 2000 to 2019) about the possible relationship between asthma and cataract. It was found that during the onset of asthma and cataract, Rho/Rock signaling pathway, Notch signaling pathway, Wnt/ß-catenin signaling pathway, PI3K/AKT signaling pathway, JAK/STAT signaling pathway, MAPK signaling pathway, TGF-ß1/Smad signaling pathway and NF-κB signaling pathway are all active, so they may have a certain correlation in pathogenesis. Asthma may be associated with cataract through the eight signaling pathways, causing inflammation or immune imbalance based on allergy that can lead to cataract. According to these studies, we speculated that the three most likely signaling pathways are PI3K/AKT, MAPK and NF-κB signaling pathway.

Structural optimization and anti-allergic activity of nucleotide aptamers target to Cε3-Cε4.

Nucleic acid aptamers have shown a broad application prospect in basic research, clinical diagnosis and treatment, new drug development and various other fields. We have screened the DNA aptamer A1 and A2 target at Cε3-Cε4 with high affinity and specificity, another aptamer A8, no affinity with Cε3-Cε4 protein, was as a negative control in this study. The structures of aptamer A1 and A2 were optimized using the deletion method, complementary sequence method, and point mutation method, to make them perform biological functions better, improve the pertinence of the subsequent modification and study the mechanism of action of aptamers coupled Cε3-Cε4. Additionally, the affinity was detected using competitive ELISA, then the most optimal and minimalist aptamer G39-A1-29C was obtained. The results indicated that the G39-A1-29C can significantly inhibit the IgE-dependent cell degranulation, but no effect in IgE-independent manner, and have a notable therapeutic effect with dose-dependent on PCA experiments in vivo. Moreover, it is found that the aptamer maintains the secondary structure through the fixed sequence, consecutive four GC pairings can significantly increase the binding affinity, and the G base on the loop region of A1 may be the key sites for binding to the domain of the target protein Cε3-Cε4. Therefore, the stem-loop structure of A1 is the structural basis of its binding, too short sequence cannot maintain the secondary structure, so that its affinity is significantly reduced. The results facilitated the modification and chemical synthesis of aptamers in next work, which provided the foundation for the development of new drugs for the treatment of allergy diseases.

[Molecular cloning and preliminary analysis of TaNOA in common wheat].

Nitric oxide (NO) is an important signaling molecule with diverse physiological functions in both animal and plant cells. In this work, we isolated the full-length cDNA and genomic DNA sequences of TaNOA-B1 encoding a putative NO associated (NOA) factor in common wheat. Bioinformatic analysis showed that TaNOA-B1 possessed a similar intron/exon structure as its orthologous genes in Arabidopsis and rice. The amino acid sequence deduced from TaNOA-B1 was more than 60% identitical to those of Arabidopsis and rice NOA1 proteins. The primary structure of TaNOA-B1 contained the zinc finger and P-loop GTPase motifs conserved in Arabidopsis and rice NOA1 proteins. There existed at least three NOA gene members in common wheat, which were mapped to homoeologous group six chromosomes 6A, 6B and 6D, respectively. TaNOA-B1 investigated in this work was located on chromosome 6B. The transcripts of TaNOA members were found mainly in leaves. TaNOA-B1-GFP fusion protein may be located in mitochondria. TaNOA transcript level was up-regulated by abscisic acid (ABA) or NaC1 treatments, indicating that TaNOA might be involved in wheat responses to abiotic stresses.