Development of a Monoclonal Antibody-Based Indirect Competitive Enzyme-Linked Immunosorbent Assay for the Rapid Detection of Gallic Acid.

Gallic acid (GA) is closely related to the quality of herbal medicines and other agricultural products. In order to facilitate the rapid detection of GA, we developed a monoclonal antibody-based ic-ELISA method. Antigens with and without connecting arms were prepared. It was found that the introduction of connecting arms (linear carbon chain) was beneficial for immune response. By utilizing hybridoma technology, a specific mAb (anti-GA-M702) was screened and identified, which exhibited a 1:40,500 antibody titer and IgG2b antibody subtype. The ic-ELISA assay was established based on anti-GA-M702. The optimal working concentrations of the encapsulated antigen and antibody were 0.5 µg/mL and 0.67 µg/mL, respectively. The ic-ELISA method showed a linear detection range of 297.17-2426.61 ng/mL for GA with a sensitivity of 849.18 ng/mL. It displayed a good applicability for the determination of GA in Galla chinensis. In conclusion, the ic-ELISA method provides an efficient approach to the rapid detection of GA in products.

Enhancement of the Physical and Functional Properties of Chitosan Films by Incorporating Galla chinensis Extract.

Composite films based on chitosan (CS) incorporating Galla chinensis extract (GCNE) at different CS/GCNE weight ratios, which are both biodegradable and multifunctional, were fabricated using the solution-casting method. The FTIR analyses indicated that a good interaction was presented among the GCNE and CS through an intermolecular hydrogen bond. The incorporation of the GCNE improved the films' elongation at break, UV-light blocking, and decreased the moisture regain (from 16.68% to 10.69%) and water absorption (from 80.65% to 54.74%). Moreover, the CS/GCNE films exhibited a strong antioxidant activity (from 57.11% to 70.37% of DPPH and from 35.53% to 46.73% of ABTS scavenging activities) mainly due to the high content of phenolic compounds in the incorporated GCNE. The CS/GCNE film-forming solution coatings demonstrated their effectiveness in preserving the quality of postharvest mangoes, specifically by minimizing the change in the firmness, weight loss, titratable acidity, and total phenolic and ascorbic acids. These findings suggest that the multifunctional composite films possess a high application potential to preserve postharvest fruits.

Methylation of hypoxia-inducible factor 3 subunit alpha contributes to poor prognosis in lung adenocarcinoma.

Hypoxia-inducible factor 3 subunit alpha (HIF3A) has been implicated in various types of cancers, while its precise role in the lung adenocarcinoma remains unclear. Our study aimed to investigate the roles of HIF3A in lung adenocarcinoma and its regulation by DNA methylation. We utilized bioinformatic tools, including UALCAN and KMPlot, to analyze the relationship between HIF3A expression, DNA methylation, and patient survival rate in lung adenocarcinoma. We also used siRNA-mediated knockdown of HIF3A and DNA-methyltransferase 1 (DNMT1), as well as the treatment of DNA methylation inhibitor 5-Azacytidine, in A549 and H1299 lung adenocarcinoma cell lines. qPCR, MTT, and cell counting assays were performed to evaluate the mRNA expression and cell viability. The bioinformatic analysis revealed that HIF3A expression was downregulated and its methylation was upregulated in lung tumor tissues. Additionally, Kaplan-Meier analysis indicated a correlation between low HIF3A expression and patient poor survival rate. We found that DNMT1 regulated HIF3A methylation. Knockdown of HIF3A promoted cancer cell proliferation. These data suggest that downregulation of HIF3A promotes tumor cell proliferation, and support that HIF3A methylation may serve as a prognostic factor for lung adenocarcinoma.

Siraitia grosvenorii (Swingle) C. Jeffrey: Research Progress of Its Active Components, Pharmacological Effects, and Extraction Methods.

Siraitia grosvenorii (Swingle) C. Jeffrey, a perennial vine of the Cucurbitaceae family, is a unique medicine food homology species from China. S. grosvenorii can be used as a natural sweetener in the food industry and as a traditional medicine for moistening the lungs, quenching a cough, smoothing the intestines, and relieving constipation. Additionally, the fruits, roots, stems, and leaves of S. grosvenorii are rich in active ingredients, and have pharmacological effects such as immune regulation, hypoglycemia, and antioxidant, hepatoprotective, and antitumor effects, etc. Therefore, S. grosvenorii has broad application prospects in the pharmaceutical industry. This paper reviews the bioactive components, pharmacological effects, and extraction methods of S. grosvenorii, summarizes them, and proposes their future development directions. This current overview highlights the value of S. grosvenorii. By documenting the comprehensive information of S. grosvenorii, the review aims to provide the appropriate guidelines for its future in-depth development and the utilization of S. grosvenorii resources for their roles as active ingredient (triterpenoids, flavonoids, and polysaccharides, etc.) sources in the food industry and in the development of functional foods.

RNF31 represses cell progression and immune evasion via YAP/PD-L1 suppression in triple negative breast Cancer.

BACKGROUND: Recently genome-based studies revealed that the abnormality of Hippo signaling is pervasive in TNBC and played important role in cancer progression. RING finger protein 31 (RNF31) comes to RING family E3 ubiquitin ligase. Our previously published studies have revealed RNF31 is elevated in ER positive breast cancer via activating estrogen signaling and suppressing P53 pathway. METHODS: We used several TNBC cell lines and xenograft models and performed immuno-blots, QPCR, in vivo studies to investigate the function of RNF31 in TNBC progression. RESULT: Here, we demonstrate that RNF31 plays tumor suppressive function in triple negative breast cancer (TNBC). RNF31 depletion increased TNBC cell proliferation and migration in vitro and in vitro. RNF31 depletion in TNBC coupled with global genomic expression profiling indicated Hippo signaling could be the potential target for RNF31 to exert its function. Further data showed that RNF31 depletion could increase the level of YAP protein, and Hippo signaling target genes expression in several TNBC cell lines, while clinical data illustrated that RNF31 expression correlated with longer relapse-free survival in TNBC patients and reversely correlated with YAP protein level. The molecular biology assays implicated that RNF31 could associate with YAP protein, facilitate YAP poly-ubiquitination and degradation at YAP K76 sites. Interestingly, RNF31 could also repress PDL1 expression and sensitive TNBC immunotherapy via inhibiting Hippo/YAP/PDL1 axis. CONCLUSIONS: Our study revealed the multi-faced function of RNF31 in different subtypes of breast malignancies, while activation RNF31 could be a plausible strategy for TNBC therapeutics.

Hypoxia-induced antizyme inhibitors 2 regulates cisplatin resistance through epithelia-mesenchymal transition pathway in non-small cell lung cancer.

Antizyme inhibitors 2 (AZIN2) was found to be associated with poor prognosis of patients with rectal cancer. However, no studies have reported whether AZIN2 functions in non-small cell lung cancer (NSCLC). This study aimed to investigate the role of AZIN2 in cisplatin (DDP) resistance in NSCLC. We established DDP resistant A549 and H1299 cell lines. The transcriptional and translational expression levels were examined using quantitative real-time polymerase chain reaction and western blot. Cell apoptosis was evaluated by caspase-3 activity and nucleosome ELISA assays. Luciferase reporter assay was employed to evaluate the impact of hypoxia-inducible factor (HIF-1α) on AZIN2 transcription. AZIN2 expression was found to be associated with DDP resistance and poor prognosis in patients with NSCLC. AZIN2 overexpression promoted cell viability, colony formation, and reduced cell apoptosis in H1299 cells and A549 upon DDP treatment. Correspondingly, AZIN2 knockdown significantly inhibited cell viability and colony formation, and increased cell apoptosis upon DDP treatment. Interestingly, AZIN2 expression in NSCLC cells was significantly induced by hypoxia condition. The occupancy of HIF-1α, an important regulator of the hypoxia response, remarkably enriched at the promoter region of AZIN2 under hypoxia condition. In addition, AZIN2 overexpression resulted in epithelial-mesenchymal transition (EMT). The results suggested that hypoxia-induced AZIN2 high expression may contribute to DDP resistance development by promoting the EMT.

Ericoid mycorrhizal fungus enhances microcutting rooting of Rhododendron fortunei and subsequent growth.

Adventitious root (AR) formation is a unique feature of plant reproduction and plays a vital role in crop production as many horticultural and forestry plants are propagated through cuttings. A growing number of reports have shown that microbes, particularly mycorrhizal fungi are able to promote AR formation, but the underlying mechanisms remain largely unclear. This study established an in vitro culture system and investigated AR formation in microcuttings of Rhododendron fortunei Lindl. inoculated with Oidiodendron maius Barron Om19, an ericoid mycorrhizal fungus strain. Hormones and precursors involved in the biosynthesis of indole-3-acetic acid (IAA) in Om19 mycelium were analyzed. Om19 was able to produce a large quantity of tryptophan (Trp) and also indole-3-pyruvate (IPA) and IAA, indicating that IAA biosynthesis in Om19 could be through a Trp-dependent pathway. After inoculation of Om19, ARs were quickly formed in microcuttings. Symbiosis related genes were activated in ARs, and Om19 effectively colonized the roots. YUC3, a key gene in plant biosynthesis of IAA and genes involved in nitrogen (N) uptake and metabolism, phosphorus (P) uptake were highly upregulated. Plants absorbed significantly greater quantity of mineral nutrients, and their growth was substantially enhanced compared to the control plants without Om19 inoculation. A working model for Om19 enhanced AR formation was proposed. The rapid formation of ARs in cuttings could be due in part to the induction of IAA biosynthesized by Om19 and also attributed to Trp catalyzed biosynthesis of IAA in plants. AR formation, in turn, provided Om19 preferred sites for colonization. Our study suggested that in addition to promoting AR formation, Om19 could potentially be used as a new biofertilizer for enhancing production of ericaceous plants, such as blueberry, cranberry, and rhododendron.

MoIVD-Mediated Leucine Catabolism Is Required for Vegetative Growth, Conidiation and Full Virulence of the Rice Blast Fungus Magnaporthe oryzae.

Isovaleryl-CoA dehydrogenase (IVD), a member of the acyl-CoA dehydrogenase (ACAD) family, is a key enzyme catalyzing the conversion of isovaleryl-CoA to ß-methylcrotonyl-CoA in the third reaction of the leucine catabolism pathway and simultaneously transfers electrons to the electron-transferring flavoprotein (ETF) for ATP synthesis. We previously identified the ETF ortholog in rice blast fungus Magnaporthe oryzae (MoETF) and showed that MoETF was essential for fungal growth, conidiation and pathogenicity. To further investigate the biological function of electron-transferring proteins and clarify the role of leucine catabolism in growth and pathogenesis, we characterized MoIVD (M. oryzae isovaleryl-CoA dehydrogenase). MoIvd is highly conserved in fungi and its expression was highly induced by leucine. The Δmoivd mutants showed reduced growth, decreased conidiation and compromised pathogenicity, while the conidial germination and appressorial formation appeared normal. Consistent with a block in leucine degradation, the Δmoivd mutants accumulated isovaleric acid, grew more slowly, fully lacked pigmentation and completely failed to produce conidia on leucine-rich medium. These defects were largely rescued by raising the extracellular pH, suggesting that the accumulation of isovaleric acid contributes to the growth and conidiation defects. However, the reduced virulence of the mutants was probably due to their inability to overcome oxidative stress, since a large amount of ROS (reactive oxygen species) accumulated in infected host cell. In addition, MoIvd is localized to mitochondria and interacted with its electron receptor MoEtfb, the ß subunit of MoEtf. Taken together, our results suggest that MoIVD functions in leucine catabolism and is required for the vegetative growth, conidiation and full virulence of M. oryzae, providing the first evidence for IVD-mediated leucine catabolism in the development and pathogenesis of plant fungal pathogens.

MiR-181a inhibits non-small cell lung cancer cell proliferation by targeting CDK1.

BACKGROUND: MicroRNAs (miRNAs) emerge as important regulators involved in malignant progression in some tumors. MiR-181a has been found to function as a tumor suppressor in some tumors including non-small cell lung cancer (NSCLC). However, the functional role of miR-181a in NSCLC still needed to be investigated. METHODS: The expression of miR-181a were determined by qRT-PCR, the association between miR-181a and clinicopathological data were performed by chi-square test and survival analysis were evaluated by Kaplan-Meier curve and log rank test. Cell proliferation and invasion were assessed by CCK8, cell colony formation and transwell assays. Luciferase reporter assay demonstrated that CDK1 was a target of miR-181a. Western blot assay detected the relative protein expression. RESULTS: In the study, our results showed that miR-181a was significantly down-regulated in non-small cell lung cancer (NSCLC) tissues and cell lines. MiR-181 expression levels were significantly associated with histological grade, N status and TNM stage in the patients and lower miR-181a predicted a poor prognosis in NSCLC patients. Furthermore, upregulation of miR-181a significantly suppressed the NSCLC cell proliferation, colony formation, and cell invasion capacities. Moreover, upregulation of miR-181a inhibited CyclinB1 and CyclinD1 expression in NSCLC cells. Luciferase activity assay results demonstrated CDK1 was a direct target of miR-181a and miR-181a inhibited cell proliferation by regulating the mRNA and protein levels of CDK1 in NSCLC cells. CONCLUSION: These data suggested that miR-181a plays a tumor suppressor and may be a potential therapeutic target for NSCLC patients.

[Comparative study of intertrochanteric fractures treated with proximal femur locking compress plate in aged].

OBJECTIVE: To compare the clinic outcomes of dynamic hip screw (DHS), intramedullary fixation (IF) and proximal femur locking plate (PF-LCP) in the treatment of intertrochanteric fractures in the elderly patients. METHODS: From July 2000 to August 2009, 165 old patients with intertrochanteric fractures were treated respectively by DHS, IF, PF-LCP. Fifty-eight patients were in DHS group including 30 males and 28 females with an average age of 71 years old; there were 30 cases of type II fracture of Jensen, 28 cases of type III fracture. Sixty-five patients were in IF group including 35 males and 30 females with an average age of 73 years old; there were 37 cases of type II fracture of Jensen, 28 cases of type III fracture. Forty-two patients were in PF-LCP group including 23 males and 19 females with an average age of 74 years old; there were 22 cases of type II fracture of Jensen,20 cases of type III fracture. The operative procedures,complications and therapeutic effects were compared among 3 groups. RESULTS: All patients were followed up from 15 to 21 months (averaged 18.3 months). The incision length and the operation time of IF group were shorter than that of DHS and PF-LCP, but there were no significant difference between DHS group and PF-LCP group. The intraoperattive blood loss, rehabilitation and healing time of IF and PF-LCP were less or shorter than that of DHS group, but there were no significant difference between IF group and PF-LCP group. The functional recovery of IF group and PF-LCP were better than that of DHS group, there were significant difference among 3 groups. The complications of PF-LCP group was fewer than that of IF group and DHS group. CONCLUSION: PF-LCP is the credible method for intertrochanteric fractures in the elderly patients, especially for severe comminuted fracture and osteoporosis, for it can reduce operation complications and benefit for fracture healing and hip functional recovery.

[Segmental root palsy (SRP) after cervical decompression surgery].

OBJECTIVE: To explore the occurrence, prevention and cure prognosis of segment root polsy (especially in C5 palsy) after cervical decompression surgery. METHODS: From February 2006 and December 2008,162 patients were operated with cervical decompression through approach for anterior or posterior in our hospital. Among them, 10 cases occurred SRP after operation included 6 males and 4 females aged from 40 to 68 with an average of 53 years old. These cases were treated with dehydration, trophic nerve, hyperbaric oxygenation. The clinical data were retrospective analzed. RESULTS: Ten patients were followed up from 8 months to 3 years with an average of 2.4 years. It was observed that all the patients recovered during a period of 4.4 months on average (ranging from 3 weeks to 8 months). CONCLUSION: SRP(especially in C5 palsy) is one of the common complications of anterior or posterior cervical decompression surgery. SRP is usually the result of various nosogenesis. As there was no effective treatment, conservative treatment is usually adopted with optimistic prognosis.

Correlation of 4-month infant feeding modes with their growth and iron status in Beijing.

BACKGROUND: Growth and development of infants has been an important topic in pediatrics for a long time. Infants must be provided with food containing all necessary nutrients. Breast milk is believed to be the most desirable natural and cheapest food for well-balanced nutrition. But with the progress in the development of substitute food in developed countries, it is thought that formula milk can meet the requirement for infant growth. During early infancy, growth, as the most sensitive index of health, is therefore a critical component in evaluating the adequacy of breast-feeding, mixed-feeding and formula feeding. Iron status is another important index of infant health. Iron deficiency anemia remains the most prevalent nutritional deficiency index in infants worldwide. This study is to compare infants in Beijing at 4 months who are on three different feeding modes (breast feeding, mixed feeding and formula feeding) in physical changes and iron status. The results may provide new mothers with support in feeding mode selection, which will also be helpful to the China Nutrition Association in feeding mode education. METHODS: This is a cohort study. One thousand and one normal Beijing infants were followed regularly for 12 months. Body weight and horizontal length were measured. Hemoglobin, red blood cell counts, mean corpuscular volume, mean corpuscular hemoglobin and serum iron were analyzed at 4 months. RESULTS: The breast feeding percentage in the first 4 months was 47.9%. The feeding mode was not significantly related to maternal delivery age, education, labor pathway nor infant sex (P>0.05). Infant boys and girls exclusively breast-fed from 0 to 4 months had the highest weight at 0-6 months. The anemia rate of breast-fed infant boys at 4 months was the highest. CONCLUSIONS: Breast feeding should be given more emphasis. It is compulsory for new mothers to breast-feed their infants if possible. Social environment should also guarantee the requirement for breast feeding. Furthermore the normal values of hemoglobin, mean corpuscular volume and serum iron, which were originally used to judge children's iron deficiency anemia, might not be optimal for evaluating infants. There might be a need to develop sex-specific cutoff levels of hemoglobin, mean corpuscular volume and serum iron for infants.