NLRP3 activation in macrophages promotes acute intestinal injury in neonatal necrotizing enterocolitis.

BACKGROUND: Macrophages are involved in various immune inflammatory disease conditions. This study aimed to investigate the role and mechanism of macrophages in regulating acute intestinal injury in neonatal necrotizing enterocolitis (NEC). METHODS: CD68, nucleotide-binding oligomerization domain, leucine-rich repeat, and pyrin domain-containing 3 (NLRP3), cysteine aspartate-specific protease-1 (caspase-1), and interleukin-1ß (IL-1ß) in paraffin sections of intestinal tissues from NEC and control patients were detected with immunohistochemistry, immunofluorescence, and western blot. Hypertonic pet milk, hypoxia and cold stimulation were used to establish a mouse (wild type and Nlrp3-/-) model of NEC. The mouse macrophage (RAW 264.7) and rat intestinal epithelial cell-6 lines were also cultured followed by various treatments. Macrophages, intestinal epithelial cell injuries, and IL-1ß release were determined. RESULTS: Compared to the gut "healthy" patients, the intestinal lamina propria of NEC patients had high macrophage infiltration and high NLRP3, caspase-1, and IL-1ß levels. Furthermore, in vivo, the survival rate of Nlrp3-/- NEC mice was dramatically improved, the proportion of intestinal macrophages was reduced, and intestinal injury was decreased compared to those of wild-type NEC mice. NLRP3, caspase-1, and IL-1ß derived from macrophages or supernatant from cocultures of macrophages and intestinal epithelial cells also caused intestinal epithelial cell injuries. CONCLUSIONS: Macrophage activation may be essential for NEC development. NLRP3/caspase-1/IL-1ß cellular signals derived from macrophages may be the underlying mechanism of NEC development, and all these may be therapeutic targets for developing treatments for NEC.

Study on fluvoxamine maleate sustained-release pellets and its compression technology / 药学学报

In this study, fluvoxamine maleate sustained-release pellet system tablets were prepared and were used to evaluate their release behaviors in vitro. Fluvoxamine maleate pellets were prepared using centrifugal-spherization method and coated by fluidized bed as bottom-spray. The multi-unit sustained-release pellets and appropriate excipients for prescription volumes were mixed uniformly and then compressed to tablets. Screening and determining the optimal formulation of drug loaded pellets through L8 (24) Taguchi experiment. Using Minitab software to design a DOE experiment with 24 partial factors, including material temperature, fan speed, atomization pressure, and spray rate to optimize the bottom spray coating process. Taking monostearate glycerol ester with a particle size of 24-40 mesh as the main diluent for tableting to relieve the delamination phenomenon between pellets and excipients during tablet pressing and reduce mechanical damage to the coating film. By examining the powder fluidity indexes such as angle of repose, bulk density, tapped density, and Hausner ratio of mixed particles, it was found that the flowability and compressibility are good and suitable for direct compression. Evaluate the basic properties of the sustained-release tablets, investigate the in vitro release behavior and study the release mechanism. The results of in vitro release test showed that the self-made sustained-release tablets could disintegrate into independent pellet units in phosphate buffer at pH 6.8 and release slowly within 24 h, which conformed to the first-order drug release model. The fluvoxamine maleate sustained-release pellet system tablets meet the requirements of preparation design and has a great commercial prospect.

Impact of COVID-19 on dietary intake, sleeping patterns and physical activity levels among Malaysian University students.

OBJECTIVES: The COVID-19 pandemic has brought a significant impact on people's behaviour and lifestyles. Limited research has been conducted on the impact of COVID-19 on Malaysian university students' lifestyle modifications. This study aims to identify the impact of COVID-19 on dietary intake, sleeping patterns and physical activity levels among Malaysian university students. METHODS: A total of 261 university students were recruited. Sociodemographic and anthropometric data were collected. Dietary intake was assessed using PLifeCOVID-19 questionnaire, sleep quality was assessed using Pittsburgh Sleep Quality Index Questionnaire (PSQI) while physical activity level was assessed using International Physical Activity Questionnaire-Short Forms (IPAQ-SF). SPSS was used to perform statistical analysis. RESULTS: 30.7% of the participants adhered to the unhealthy dietary pattern, 48.7% had poor quality of sleep and 59.4% engaged in low physical activity levels during the pandemic. Unhealthy dietary pattern was significantly associated with a lower IPAQ category (p=0.013), and increased time spent sitting (p=0.027) during the pandemic. Participants being underweight before the pandemic (aOR=2.472, 95% CI=1.358-4.499), increased takeaway meal consumption (aOR=1.899, 95% CI=1.042-3.461), increased snacking between meals consumption (aOR=2.989, 95% CI=1.653-5.404) and engaged in a low level of physical activity during pandemic (aOR=1.935, 95% CI=1.028-3.643) were the predictors of unhealthy dietary pattern. CONCLUSIONS: The university students' dietary intake, sleeping patterns, and physical activity levels were impacted in different ways during the pandemic. Strategies and interventions should be developed and implemented to improve the dietary intake and lifestyle of the students.

Optimal design and application of information management system of biobank in hospitals / 中华医学科研管理杂志

Objective:To establish a standardized information management system (IMS) for preserving, managing, querying, and performing statistics on biospecimens and their clinical data, which is conducive to improving the utilization of biobank.Methods:Under the premise of ensuring operating environment and data security, a database-based data logic relationship model is created and applied to the IMS to manage and analyze biospecimens and their supporting clinical information of patients enrolled in the biobank of our center.Results:To ensure the establishment of the follow-up cohort, biospecimens and clinical information of inpatients and outpatients were continuously collected in the biobank of our center. Since December 2014, more than 270 000 biospecimens from inpatient, outpatient, and scientific research have been preserved. The IMS optimized by this model efficiently completes the basic work of the biobank. At the same time, the data can be queried jointly and in batches, and then converted into a report format for statistical analysis.Conclusions:The IMS of our center is suitable for application and popularization as a construction and management model for the hospital-level biobank, which meets the daily work of the biobank and diverse research needs, and provides a convenient platform and rich resources for the development of precision medicine.

Gan-Qing-Ning Formula Inhibits the Growth of Hepatocellular Carcinoma by Promoting Apoptosis and Inhibiting Angiogenesis in H22 Tumor-Bearing Mice.

OBJECTIVE: Gang-Qing-Ning (GQN) is a traditional Chinese medicine formula that has been used in the treatment of hepatocellular carcinoma (HCC) in the folk population for decades. However, scientific validation is still necessary to lend credibility to the traditional use of GQN against HCC. This study investigates the antitumor effect of GQN on H22 tumor-bearing mice and its possible mechanism. METHODS: Fifty H22 tumor-bearing mice were randomly assigned to five groups. Three groups were treated with high, medium, and low dosages of GQN (27.68, 13.84, and 6.92 g/kg, respectively); the positive control group was treated with cytoxan (CTX) (20 mg/kg) and the model group was treated with normal saline. After 10 days' treatment, the tumor inhibitory rates were calculated. Pathological changes in tumor tissue were observed, and the key proteins and genes of the mitochondrial apoptosis pathway were measured, as well as the mRNA expression levels of VEGF in tumor tissue. RESULTS: The tumor inhibitory rates of high, medium, and low dosages of GQN groups were 47.39%, 38.26%, and 22.17%, respectively. The high dosage of the GQN group significantly increased the protein and mRNA expression levels of Bax, Cyt-C, and cleaved Caspase 3 (or Caspase 3) (P < 0.01) but decreased the expression levels of Bcl-2, VEGF, and microvessel density (MVD) (P < 0.01). CONCLUSIONS: The high dosage of GQN can significantly inhibit the tumor growth in H22 tumor-bearing mice. It exerts the antitumor effect by enhancing proapoptotic factors and inhibiting the antiapoptotic factor of the mitochondrial apoptosis pathway and inhibiting tumor angiogenesis.

Rrp6 Regulates Heterochromatic Gene Silencing via ncRNA RUF6 Decay in Malaria Parasites.

The heterochromatin environment plays a central role in silencing genes associated with the malaria parasite's development, survival in the host, and transmission to the mosquito vector. However, the underlying mechanism regulating the dynamic chromatin structure is not understood yet. Here, we have uncovered that Plasmodium falciparum Rrp6, an orthologue of eukaryotic RNA exosome-associated RNase, controls the silencing of heterochromatic genes. PfRrp6 knockdown disrupted the singular expression of the GC-rich ncRNA RUF6 family, a known critical regulator of virulence gene expression, through the stabilization of the nascent transcripts. Mechanistic investigation showed that the accumulation of the multiple RUF6 ncRNAs triggered local chromatin remodeling in situ, which activated their adjacent var genes. Strikingly, chromatin isolation by RNA purification analysis (ChIRP-seq) revealed that a remarkable RUF6 ncRNA had interacted with distal heterochromatin regions directly and stimulated a global derepression effect on heterochromatic genes, including all variant gene families and the sexual commitment-associated regulator ap2-g gene. Collectively, Rrp6 appears to conduct the epigenetic surveillance of heterochromatic gene expression through controlling RUF6 levels, thereby securing antigenic variation and sexual commitment of malaria parasites during the infection of the host.IMPORTANCE Malaria remains a major public health and economic burden. The heterochromatin environment controls the silencing of genes associated with the fate of malaria parasites. Previous studies have demonstrated that a group of GC-rich ncRNAs (RUF6) is associated with the mutually exclusive expression of var genes, but the underlying mechanisms remain elusive. Here, through a series of genetic manipulation and genome-wide multiomics analysis, we have identified the plasmodial orthologue of RNA exosome-associated Rrp6 as an upstream regulator of RUF6 expression and revealed that the dysregulation of RUF6 upon Rrp6 knockdown triggered local chromatin alteration, thereby activating most heterochromatic genes via direct interaction of RUF6 and distal gene loci. This finding not only uncovered the in-depth mechanism of RUF6-mediated regulation of heterochromatic genes but also identified Rrp6 as a novel regulator of gene expression in human malaria parasites, which provides a new target for developing intervention strategies against malaria.

Orthogonal optimization of prokaryotic expression of a natural snake venom phospholipase A2 inhibitor from Sinonatrix annularis.

Phospholipase A2 (PLA2) is a calcium-dependent enzyme that is involved in inflammatory processes such as the liberation of free arachidonic acid from the membrane pool for the biosynthesis of eicosanoids. Snake venom are known containing PLA2s (svPLA2s) which exhibit a wide variety of pharmacological effects including neurotoxicity, cardiotoxicity, myotoxicity and hemorrhage. Therefore, inhibition of svPLA2 would be advantageous to successful envenomation treatment. A gamma type PLI (PLA2 inhibitor) has been extracted from the serum of Sinonatrix annularis, a non-venomous snake indigenous to China. This showed strong inhibition of Deinagkistrrodon acutus PLA2, however, the PLIγ level in the serum and snake resource are not sufficiently sustainable for further research. To overcome these limitations, we constructed a His6-PLIγ pET28 fusion expression vector and transformed Escherichia coli BL21. To improve the expression of PLIγ, an orthogonal experiment [L16(4)(5)] was performed to optimize induction parameters. The optimized condition was determined to be: induction by 0.4 mM isopropyl-ß-D-thiogalactoside (IPTG) for 6 h to the recombinant BL21 after its OD600 was 0.8, with continuous shaking cultivation at 190 rpm and 35 °C. Under these conditions, the amount of expressed protein could reach 57 mg/L. The His6-PLIγ was purified by nickel affinity chromatography and renatured by On-column refolding. The resulting PLIγ showed a good inhibitory effect of enzymatic activities to venom PLA2 isolated from D. acutus. Moreover, the PLIγ had a wide anti-hemorrhage activities to D. acutus, Naja atra and Agkistrodon halys venom.

Dynamic dialysis: an efficient technique for large-volume sample desalting.

Dialysis is a well-known technique for laboratory separation. However, its efficiency is commonly restricted by the dialyzer volume and its passive diffusion manner. In addition, the sample is likely to be precipitated and inactive during a long dialysis process. To overcome these drawbacks, a dynamic dialysis method was described and evaluated. The dynamic dialysis was performed by two peristaltic pumps working in reverse directions, in order to drive countercurrent parallel flow of sample and buffer, respectively. The efficiency and capacity of this dynamic dialysis method was evaluated by recording and statistically comparing the variation of conductance from retentate under different conditions. The dynamic method was proven to be effective in dialyzing a large-volume sample, and its efficiency changes proportionally to the flow rate of sample. To sum up, circulating the sample and the buffer creates the highest possible concentration gradient to significantly improve dialysis capacity and shorten dialysis time.

Novel microdeletion in the transforming growth factor beta type II receptor gene is associated with giant and large cell variants of nonsmall cell lung carcinoma.

Mutations in the tumor suppressor gene transforming growth factor beta (TGFB) Type II receptor (TGFBR2) are frequently found in many cancers with microsatellite instability, but are less common in lung cancer. In the present study, we looked for mutations in TGFBR2 in nonsmall cell lung carcinoma (NSCLC) cells and tissues. A novel homozygous microdeletion (c.492_507del) was identified in two cell lines derived from the same giant cell carcinoma (GCC) and was confirmed in the corresponding tumor tissues. Furthermore, a heterozygous c.492_507del was found in the germ-line of one patient, as well as in the other GCC cases and some large cell carcinomas (LCC) but not in other subtypes of NSCLC. The 16 bp-microdeletion introduced a premature stop codon at positions 590-592 of the cDNA, resulting in a truncated TGFBR2 protein with a mutated transmembrane domain and loss of kinase domain. The GCC cells were characterized as being unresponsive to TGFB induction both in growth inhibition and stimulation of extracellular matrix protein. Moreover, after the reconstitution of wild-type TGFBR2 expression, the sensitivity to TGFB was restored. Therefore, mutated TGFBR2 seems to play an important role in the abrogation of TGFB signal transduction in GCC cells.

Zinc-mediated chain extension reaction of 1,3-diketones to 1,4-diketones and diastereoselective synthesis of trans-1,2-disubstituted cyclopropanols.

[reaction: see text] A variety of 1,3-diketones can be efficiently converted into the corresponding 1,4-diketones and trans-1,2-disubstituted cyclopropanols by using organozinc species in one-pot reactions. It was found that 2.3 equiv of CF3CO2ZnCH2I was effective to give the corresponding chain-extended products in 44-85% yields, while a mixture of organozinc species formed from 4.0 equiv of Et2Zn, 2.0 equiv of CF3CO2H, and 4.0 equiv of CH2I2 resulted in the formation of trans-1,2-disubstituted cyclopropanols with quite good yields and diastereoselectivity.

Zinc-mediated ring-expansion and chain-extension reactions of beta-keto esters.

[Chemical reaction: see text] The reaction of cyclic beta-keto esters with CF3CO2ZnCH2I provided the corresponding ring-expanded products in moderate to good yields. Although alpha-substituted acyclic beta-keto esters reacted with much less efficient, chain-extension reaction of simple beta-keto esters also proceeded effectively to generate gamma-keto esters in high yields.

[Preoperational misdiagnosis of extragastrointestinal stromal tumors as ovarian cancer: report of three cases with literature review].

OBJECTIVE: This report was to review how extragastrointestinal stromal tumor (EGIST) was similar to ovarian cancer, its differential diagnosis and treatment. METHODS: Three cases of EGIST were reported and discussed by reviewing literatures. RESULTS: The cases of EGIST were presented with peritoneal and pelvic mass, or ascites, diagnosed as ovarian cancer at first, and finally diagnosed as EGIST by pathological method and positive CD(117) and CD(34) expression. Surgery in combination with chemotherapy was an effective therapeutic method. CONCLUSIONS: Clinical presentation of EGIST is similar to ovarian cancer in some aspects. Detection of CD(117) and CD(34) positive expression is helpful in the diagnosis of EGIST.

[Immunohistochemical study on telomerase activity in ovarian sex-cord stromal tumors].

BACKGROUND & OBJECTIVE: Telomerase activity has been detected in a broad range of human cancers including epithelial malignancies from in female genital tract, but the expression of the telomerase activity in ovarian sex-cord tumors has not been reported up to now. In this report we investigated the telomerase activity in ovarian sex-cord stromal tumors and its relationship with clinicopathological feature. METHODS: Twenty-five cases of ovarian sex-cord stromal tumor, including granulosa cell tumors, thecofibromas, sclerosing stromal tumors, and Sertoli-Leydig cell tumors were retrieved, and the expression of human telomerase reverse transcriptase (hTRT) was assessed immunohistochemically using polyclonal antibody H231. RESULTS: hTRT were positive in 60.0% of all cases, and the signal mainly located in the cytoplasm of tumor cells, especially in luteinized cells and Ledig cells. The positive rate was significantly related to different histological types and the patients' clinical endocrinal manifestation (P value were 0.01 and 0.041, respectively), but no statistically significant difference was found between the expression of hTRT and tumor histological grade or patients' prognosis. CONCLUSIONS: In ovarian sex-cord stromal tumors, the tumor component differentiating to luteinized stromal cell has high telomerase activity, and the telomerase activity may play an important role in patients' endocrinal disorder.

Detection of hepatitis C virus NS5 protein and genome in Chinese carcinoma of the extrahepatic bile duct and its significance.

AIM:To investigate the hepatitis C virus (HCV) infection in the tissues of carcinoma of extrahepatic bile duct and study their correlation.METHODS:HCV NS5 protein and HCV RNA were detected by labeled streptavidin biotin (LSAB) method and in situ reverse transcription polymerase chain reaction(IS-RT-PCR) in sections of 51 cases of carcinoma of extrahepatic bile duct and 34 cases of control group (without malignant biliary disease).RESULTS:In 51 cases of carcinoma of extrahepatic bile duct, HCV NS5 protein was detected in 14 (27.5%), which was clearly stained in the cytoplasm of cancer cell but not in the nucleus or cell membrane. HCV RNA was detected in 18 (35.4%), which was located in the nucleus of cancer cell in 12 cases and in the cytoplasm in 6 cases. HCV NS5 protein and RNA coexistence was found in 2 cases. In 34 cases of control group, HCV RNA was detected in 2 (5.9%). HCV NS5 protein and RNA positive cells were found either scattered or in clusters.CONCLUSION:The prevalence of hepatitis C viral infection in the tissues of carcinoma of extrahepatic bile duct was significantly higher than in control group (X(2) = 9.808,P=0.002). The findingssuggest a correlation between HCV infection and carcinoma of extrahepatic bile duct, which is different from the traditional viewpoint. HCV infection might be involved in the development of carcinoma of extrahepatic bile duct.