[Experimental study on the inhibition of the NLRP3 signaling pathway with Shengsan Jiedu Huayu decoction to alleviate inflammatory injury in rats with acute-on-chronic liver failure].

Objective: To observe the therapeutic effect of Shengsan Jiedu Huayu decoction in alleviating inflammatory liver injury in rats with acute-on-chronic liver failure (ACLF) and its effect on the activation intensity for the NLRP3 signaling pathway. Methods: 63 SD rats were randomly divided into a blank group, a model group, and low-, medium-, and high-dose groups of Shengsan Jiedu Huayu decoction (7.29 g/kg/d, 14.58 g/kg/d, and 29.16 g/kg/d). The ACLF rat model was replicated using carbon tetrachloride combined with d-galactosamine and lipopolysaccharide. Different dose gradients of the Shengsan Jiedu Huayu decoction were used for a five-day intervention treatment, and then rat serum and tissue samples were collected. A biochemical analyzer was used to detect the serum levels of ALT, AST, and TBIL in rats. ELISA was used to detect serum IL-18 and IL-1ß content. HE staining was used to observe histomorphological changes in liver tissue. Immunohistochemistry was used to detect GSDMD expression in liver tissue. Western blot and PCR were used to detect NLRP3, Caspase1, ASC, TLR4, IL-1ß, IL-18 protein, and mRNA expression levels.The groups were compared using analysis of variance and the rank-sum test. Results: Compared with the blank group, the model group's rat liver tissue was severely injured. Serum levels of ALT, AST, and TBIL, inflammatory factors IL-1ß and IL-18, and the GSDMD protein expression level, NLRP3 expression level, TLR4, caspase 1, ASC, IL-1ß, IL-18 protein, and mRNA (P<0.01) were all significantly increased in the model than the blank group (P<0.01). Additionally, compared with the model group, the low-, medium-, and high-dose groups of Shengsan Jiedu Huayu decoction had improved liver tissue injury in ACLF rats, while the serum levels of ALT, AST, TBIL, IL-1ß, IL-18, liver tissue GSDMD protein, NLRP3, TLR4, caspase 1, and ASC expressions were all lower in the different dose gradients of the Shengsan Jiedu Huayu decoction than the model group, with the most evident reduction in the high-dose group (P<0.01). Conclusion: Shengsan Jiedu Huayu decoction can weaken the activation intensity of the NLRP3 signaling pathway, alleviate liver tissue pathological injury, reduce inflammatory factor release, and alleviate inflammatory liver injury in ACLF rats.

[Preliminary clinical effect evaluation of digital head and neck radiotherapy oral positioning stents].

Objective: To develop a designing software of digital oral positioning stent for radiotherapy of head and neck, and to compare its clinical effect with traditional oral positioning stents made by lost wax process. Methods: Thirty patients with nasopharyngeal cancer who received oral examination before radiotherapy in the prosthodontics department from July to December, 2021, were selected and divided into three groups according to the patients' wishes, 10 per group: one group without radiotherapy oral positioning stents, one group with traditional oral positioning stents (traditional stents group), and the third group with digital oral positioning stents (digital stents group). Patients' ages range from 20 years old to 71 years old. There were 15 males and 15 females involved in this study. The manufacturing time and comfort of the two positioning stents were evaluated, and the radiation doses of the radiotherapy target areas and surrounding healthy tissues were statistically analyzed at the end of radiotherapy. Results: The manufacturing time of digital stents group [(209±7) min] was much less than that of traditional stents group [(490±10) min] (t=69.85, P<0.001). The comfort of patients' wearing of digital stents [first wearing: 5 (3, 6) score; at the end of radiotherapy: 4 (3, 5) score] was better than that of traditional ones [first wearing: 7 (3, 7) score; at the end of radiotherapy: 7 (3, 7) score] (U=22.00, P=0.033; U=20.50, P=0.023). There was no significant differences in the target radiation doses among the three groups, and the radiation doses of tongue [traditional stents group: (36.74±5.45) Gy; digital stents group: (35.96±4.98) Gy] and mandible [traditional stents group: (35.46±4.19) Gy; digital stents group: (35.34±3.98) Gy] were significantly lower in the patients wearing oral positioning stents than in the patients without oral positioning stents [tongue: (41.49±4.46) Gy; madible: (39.32±3.52) Gy] (P<0.05). Conclusions: Oral positioning stents for nasopharyngeal carcinoma radiotherapy could greatly reduce the exposure doses of tongue and madible of patients. Digital oral positioning stents designed and manufactured by independently developed software had higher production efficiency than the traditional method, and patients' evaluation of comfort was better.

[Clinical characteristics and related factors of acute tubular necrosis in patients with minimal change disease].

Objective: To investigate the clinical characteristics and related factors of acute tubular necrosis (ATN) in patients with minimal change disease (MCD). Methods: Patients from Chinese PLA General Hospital who were pathologically diagnosed with MCD and had clinical manifestations of nephrotic syndrome from January 1, 2013 to December 31, 2019 were included. The clinical and pathological data of patients were retrospectively analyzed. Meanwhile, the incidence and clinical characteristics of ATN in different age groups were compared. The risk factors for ATN were assessed using binary logistic regression. Results: A total of 525 patients were included, with a gender ratio of 1.56∶1 (male: female), aged 33 (21, 48) years old. ATN occurred in 49 (9.3%) of 525 patients, of which 34 were male and 15 were female. The incidence of ATN increased with age in MCD patients of different age groups (χ(2)=31.442, P<0.001). The incidence of ATN in groups of age≤20 years, 21-40 years, 41-60 years, and >60 years was 2.4% (3/123), 5.2% (10/192), 13.2% (20/152) and 27.6% (16/58), respectively. Elevations of alanine aminotransferase (ALT), aspartate aminotransferase (AST), γ-glutamyl transpeptidase (GGT) and serum IgE occurred in 92 patients (17.5%), 53 patients (10.1%), 99 patients (18.9%), and 303 patients (57.7%), respectively. There were significant differences in age, ALT, serum creatinine, serum urea nitrogen, history of diabetes and history of hypertension between non-ATN group and ATN group (all P<0.05). The results of logistic regression analysis showed that>40 years old (OR=6.283, 95% CI: 2.695-14.649, P<0.001) and serum albumin (OR=0.924, 95% CI: 0.857-0.997, P=0.040) was independently associated with ATN in MCD patients. Conclusion: Age>40 years is an independent risk factor and serum albumin is a protective factor for ATN in MCD patients.

SGK2 promotes renal cancer progression via enhancing ERK 1/2 and AKT phosphorylation.

OBJECTIVE: Increasing studies reported that the serum- and glucocorticoid-inducible kinases (SGKs) contributed to the tumorigenesis of various cancer. In this article, we are aiming to explore the function of SGK2 in renal cell cancer (RCC). PATIENTS AND METHODS: In this study, the SGK2 expression was quantified by Western blot (WB) in multiple RCC cell lines. And in vitro SGK2 knockdown and overexpression experiments were also performed. In addition, molecular function analysis was performed using FunRich software V3. The Cancer Genome Atlas (TCGA) database was retrieved to verify the association between the SGK2 expression and the prognosis of RCC patients. RESULTS: We found that SGK2 was up-regulated in RCC tissues compared with adjacent normal tissues, and the SGK2 expression also increased in various RCC cell lines compared to that in the normal epithelial cell line HK-2. Meanwhile, the SGK2 expression was significantly associated with the survival rate of RCC patients. Functional experiments showed that silencing SGK2 expression inhibited RCC cells proliferation, migration, colony formation and invasion abilities in vitro, whereas opposite results were uncovered after overexpressing SGK2 in RCC cells. Furthermore, functional analyses showed that SGK2 related genes were associated with protein serine/threonine kinase activity, guanosine triphosphatase (GTPase) activity, guanyl-nucleotide exchange factor activity, and motor activity. Protein interaction analysis identified that growth factor receptor-bound protein 2 (GRB2), one of the most important upstream components in the growth factor signaling pathway, was significantly enriched in SGK2 related genes. In addition, the WB assay validated that SGK2 could promote the phosphorylation of ERK 1/2 and AKT. CONCLUSIONS: Our results suggested that SGK2 promoted RCC progression by mediating the phosphorylation of extracellular regulated protein kinases (ERK) 1/2 and Protein kinase B (AKT/PKB), indicating that SGK2 might serve as a potential prognostic marker and therapeutic target for renal cancer patients.

Changes in methylation of genomic DNA from chicken immune organs in response to H5N1 influenza virus infection.

DNA methylation is an important epigenetic modification in eukaryotes, which plays a significant role in regulating gene expression. When the host is invaded by the influenza virus, gene expression is regulated via changes in DNA methylation levels or patterns, leading to the activation or suppression of relevant signaling pathways or networks, triggering a series of immune responses against viral invasion. Here, we investigated the changes in genomic DNA methylation in the immune organs of chicken infected with H5N1 influenza virus. Genome-wide DNA methylation levels in the spleen, thymus, and bursa of Fabricius of specific pathogen-free (SPF) chicken infected with the Guangdong (G-H5N1) and Anhui (A-H5N1) H5N1 strains, and water (control) were analyzed by fluorescence-labeled methylation-sensitive amplified polymorphism (F-MSAP). The results indicated that total DNA methylation levels did not differ between spleen genomic DNA in chicken treated with different viral strains and the control (P > 0.05). However, the total DNA methylation levels were significantly upregulated in the thymus (P < 0.01) and bursa (P < 0.05) of chicken in the A-H5N1 group compared to those in the G-H5N1 and control groups. These results provide a basis for the screening of avian influenza-resistance genes or methylation markers, analyzing the epigenetic regulation mechanisms of avian influenza, and performing selective breeding for disease resistance.

The first genetic map of a synthesized allohexaploid Brassica with A, B and C genomes based on simple sequence repeat markers.

KEY MESSAGE: We present the first genetic map of an allohexaploid Brassica species, based on segregating microsatellite markers in a doubled haploid mapping population generated from a hybrid between two hexaploid parents. This study reports the first genetic map of trigenomic Brassica. A doubled haploid mapping population consisting of 189 lines was obtained via microspore culture from a hybrid H16-1 derived from a cross between two allohexaploid Brassica lines (7H170-1 and Y54-2). Simple sequence repeat primer pairs specific to the A genome (107), B genome (44) and C genome (109) were used to construct a genetic linkage map of the population. Twenty-seven linkage groups were resolved from 274 polymorphic loci on the A genome (109), B genome (49) and C genome (116) covering a total genetic distance of 3178.8 cM with an average distance between markers of 11.60 cM. This is the first genetic framework map for the artificially synthesized Brassica allohexaploids. The linkage groups represent the expected complement of chromosomes in the A, B and C genomes from the original diploid and tetraploid parents. This framework linkage map will be valuable for QTL analysis and future genetic improvement of a new allohexaploid Brassica species, and in improving our understanding of the genetic control of meiosis in new polyploids.

Optimal pollinator attraction strategies in Trollius ranunculoides Hemsl. (Ranunculaceae) at different altitudes: increased floral display or promotion of nectar output?

For alpine plant species, patterns of resource allocation to functional floral traits for pollinator attraction can be highly significant in adaptation to low pollinator abundance and consequent pollen limitation. Increased pollination can be achieved either through a larger floral display or production of more pollen rewards. In this study, variation in resource allocation to different components for pollinator attraction was studied along an altitudinal gradient in Trollius ranunculoides, an obligate self-incompatible out-crosser of the Qinghai-Tibet Plateau. We compared resource allocation to conspicuous yellow sepals (which mainly provide visual attraction) and degenerate petals (which provide the major nectar reward) between populations at four altitudes. Furthermore, we investigated the contribution of sepals and petals to pollinator attraction and female reproductive success in an experiment with sepal or petal removal at sites at different altitudes. At the level of single flowers, resource allocation increased to sepals but decreased to petals with increasing altitude. Consistent with these results, sepals contributed much more to visitation rate and seed set than petals, as confirmed in the sepal or petal removal experiment. Sepals and petals contributed to female reproductive success by ensuring visitation rate rather than visitation duration. To alleviate increasing pollen limitation with increasing altitude, resource allocation patterns of T. ranunculoides altered to favour development of sepals rather than petals. This strategy may improve pollination and reproductive success through visual attraction (sepal) rather than nectar reward (petal) over a gradient of decreasing pollinator abundance.

Size-dependent sex allocation in Aconitum gymnandrum (Ranunculaceae): physiological basis and effects of maternal family and environment.

Theory predicts size-dependent sex allocation (SDS): flowers on plants with a high-resource status should have larger investment in females than plants with a low-resource status. Through a pot experiment with Aconitum gymnandrum (Ranunculaceae) in the field, we examined the relationship between sex allocation of individual flowers and plant size for different maternal families under different environmental conditions. We also determined the physiological base of variations in plant size. Our results support the prediction of SDS, and show that female-biased allocation with plant size is consistent under different environmental conditions. Negative correlations within families showed a plastic response of sex allocation to plant size. Negative genetic correlations between sex allocation and plant size at the family level indicate a genetic cause of the SDS pattern, although genetic correlation was influenced by environmental factors. Hence, the size-dependency of sex allocation in this species had both plastic and genetic causes. Furthermore, genotypes that grew large also had higher assimilation ability, thus showing a physiological basis for SDS.

Reproduction and cytogenetic characterization of interspecific hybrids derived from crosses between Brassica carinata and B. rapa.

The tri-genomic hybrid (ABC, 2n=27) between Brassica carinata (BBCC, 2n=34) and B. rapa (AA, 2n=20) is a unique material for studying genome relationships among Brassica species and a valuable bridge for transferring desirable characteristics from one species to the other within the genus Brassica. The crossability between B. carinata and B. rapa was varied with the cultivar of B. rapa. Hybrid pollen mother cells (PMCs), confirmed by morphological observation and molecular marker assay, could be grouped into 20 classes on the basis of chromosome pairing configurations. More than 30% of the PMCs had nine or more bivalents. Genomic in situ hybridization confirmed that two of the bivalents most likely belonged to the B genome. Nearly one-half of the PMCs had trivalents (0-2) and quadrivalents (0-2), which revealed partial homology among the A, B, and C genomes and suggested that there is a good possibility to transfer genes by means of recombination among the three genomes. The advantages of using the tri-genomic hybrids as bridge material for breeding new types of B. napus are discussed.

Development of PCR-based markers linked to the fertility restorer gene for the polima cytoplasmic male sterility in rapeseed (Brassica napus L.).

Bulked segregant analysis was used to identify RAPD markers linked to the Rfp of pol cms in rapeseed (Brassica napus L.) from the fertile and sterile DNA bulks. DNA bands amplified from 1040 random 10-mer primers were screened. Two polymorphic bands S1019(720) and S1036(810) were found linked to the Rfp locus at the same side with a map distance of 5.8 cM and 12.3 cM respectively. These polymorphic fragments were cloned and sequenced. Nucleotide sequence information was used to design 20-24-mer oligo nucleotide primers for PCR amplification. The SCAR markers that generated from the long primers showed the same pattern of segregation as the original RAPD markers in the backcross population. The SCAR markers would facilitate selection on the Pol CMS restorer lines in rapeseed.