Anemia among rural children aged 3-6 in central China and its association with dietary behaviors / 中国学校卫生

Objective@#To understand the status of anemia among rural children aged 3-6 in central China and its relationship with dietary behavior, and to provide reference for prevention and intervention of anemia in rural children.@*Methods@#From April to June 2021, stratified cluster sampling was used to select 1 246 preschool children aged 3 to 6 years old in rural areas in central China. Peripheral blood of the ring finger was collected. Dietary behaviors regarding consumption of cereals and potatoes, fruits, vegetables, soy products, milk, animal foods, food diversity, and drinking water were investigated. Logistic regression analysis was used to analyze the association between anemia and dietary behaviors.@*Results@#The detection rate of anemia in rural children aged 3-6 in central China was 13.88%. The anemia detection rate of left behind children (19.00%) was higher than that of non left behind children (11.27%), and the difference was statistically significant ( χ 2=13.93, P <0.01). Logistic regression analysis showed that the proportion of left behind children ( OR=1.28, 95%CI =1.16-1.56) with anemia was higher than that of non left behind children ( P <0.01). Consumption of animal food intake ≥3 times/week ( OR=0.82, 95%CI =0.69-0.91), and ≥2 kinds of vegetables ( OR= 0.86, 95%CI =0.71-0.93) were associated with lower rate of anemia ( P <0.05).@*Conclusion@#The detection rate of anemia in rural children aged 3-6 years in central China is relatively high, especially in left behind children. Dietary literacy of caregivers should be further improved regarding increased intake of animal foods and vegetables to reduce the risk for anemia in preschoolers.

Alterations in the expression level of visfatin in the lungs of piglets infected with PRRSV and its effect on PRRSV replication.

Porcine reproductive and respiratory syndrome (PRRS) is a highly contagious disease caused by PRRS virus (PRRSV), characterized by sow reproductive failure and respiratory symptoms in pigs of all ages. PRRSV mainly causes severe lung damage by invading alveolar macrophages. Visfatin is closely related to acute lung injury, immune response and inflammation along with virus invasion to the host. Therefore, the current study was performed to clarify the relationship between visfatin and PRRSV infection. We used ternary piglets to construct a piglet model to explore the expression of visfatin and tight junction protein in lung injury induced by PRRSV infection, and then further studied the inhibition effect of visfatin on PRRSV replication by PRRSV infection of Marc-145 cells. Our results indicated that both PRRSV attenuated and virulent infections could damage the lung tissues, which could not only lead to severe inflammatory reaction (such as increased expression of TNF-α, TGF-ß, IL-8 and IL-10) in lung tissues of piglets, but also brought about the sharp decrease of ZO-1 and Tricellulin expressions resulting in impaired alveolar epithelial barrier. Meanwhile, we found significantly up-regulated expression of visfatin in lungs and serum of pigs after PRRSV infection that were related to both the degree of lung injury and the virulence of PRRSV strain. Moreover, visfatin might inhibit the PRRSV infection to Marc-145 cells in time dependent fashion. Hence, the current investigation provides the novel information about the effect of visfatin and PRRSV co-culture on Marc-145 cells and the effect of visfatin on PRRSV proliferation at different time points.

Development of sandwich Enzyme-Linked Immunosorbent Assay for the detection of porcine epidemic diarrhea virus in fecal samples.

Porcine epidemic diarrhea virus (PEDV) causes acute diarrhea and dehydration in new-born piglets with subsequent economic losses to swine industry. In the current study, gene encoding of 381aa-792aa spike protein (S1) with the main epitope relative to virus neutralization of PEDV was amplified by RT-PCR and inserted into vector pET-30A(+). The plasmid was transferred into Escherichia coli BL21 (DE3). Meanwhile, recombinant protein expression was induced by isopropy1-ß-galactopyranoside (IPTG). After denaturation and renaturation of inclusion bodies, the S1 protein was obtained by using purified recombinant S1 protein in immunized female BALB/c mice. Monoclonal antibodies (MAb) against S1 protein, named 4C7 by hybridoma technique were gained successfully. The result showed that MAb can specifically respond to S1 protein and PEDV via ELISA, Western bolt and immunofluorescence assay methods. A sandwich ELISA (S-ELISA) was established by using the captured monoclonal antibodies 4C7. The sensitivity and specificity were compared between S-ELISA and RT-PCR, which showed similar sensitivity and specificity. This work indicated that S-ELISA would be a significant tool alongside a specific diagnostic reagent for PEDV in future.

Regulation of glycolytic metabolism by autophagy in liver cancer involves selective autophagic degradation of HK2 (hexokinase 2).

Impaired macroautophagy/autophagy and high levels of glycolysis are prevalent in liver cancer. However, it remains unknown whether there is a regulatory relationship between autophagy and glycolytic metabolism. In this study, by utilizing cancer cells with basal or impaired autophagic flux, we demonstrated that glycolytic activity is negatively correlated with autophagy level. The autophagic degradation of HK2 (hexokinase 2), a crucial glycolytic enzyme catalyzing the conversion of glucose to glucose-6-phosphate, was found to be involved in the regulation of glycolysis by autophagy. The Lys63-linked ubiquitination of HK2 catalyzed by the E3 ligase TRAF6 was critical for the subsequent recognition of HK2 by the autophagy receptor protein SQSTM1/p62 for the process of selective autophagic degradation. In a tissue microarray of human liver cancer, the combination of high HK2 expression and high SQSTM1 expression was shown to have biological and prognostic significance. Furthermore, 3-BrPA, a pyruvate analog targeting HK2, significantly decreased the growth of autophagy-impaired tumors in vitro and in vivo (p < 0.05). By demonstrating the regulation of glycolysis by autophagy through the TRAF6- and SQSTM1-mediated ubiquitination system, our study may open an avenue for developing a glycolysis-targeting therapeutic intervention for treatment of autophagy-impaired liver cancer.

Effects of Boron Supplementation on Expression of Hsp70 in the Spleen of African Ostrich.

Increased synthesis of heat shock protein 70 (Hsp70) occurs in prokaryotes and eukaryotes in response to physiological, environmental, and chemical exposures, thus allowing the cell survival from fatal conditions. Hsp70 cytoprotective properties may be clarified by its anti-apoptotic function. Boron has been reported to play an essential role in various organ developments and metabolisms. However, it is not known if boron is also able to modulate the Hsp70. In the present study, the actions of boron on ostrich spleen and expression level of Hsp70 were investigated. Thirty healthy ostrich chicks were randomly assigned to six groups: groups I, II, III, IV, V, and VI and fed the basal diet spiked with 0-, 40-, 80-, 160-, 320-, and 640-mg boric acid (BA)/L, respectively, in drinking water. The histomorphological examination in the spleen was done by hematoxylin and eosin (HE) staining. The expression level of Hsp70 was analyzed by immunohistochemistry (IHC) and western blotting, and mRNA expression of Hsp70 was investigated by quantitative real-time PCR (qPCR). In order to investigate apoptosis, TUNEL assay reaction in all treatment groups was analyzed. Our results showed that the histological structure of spleen up to 160 mg/L BA supplementation groups well developed. The Hsp70 expression level first induced at low-dose groups (up to group IV) and then inhibited dramatically in high-dose groups (V and VI) while comparing with the group I (0 mg BA). The TUNEL assay reaction revealed that the cell apoptosis amount was decreased in group IV, but in group V and especially in group VI, it was significantly increased (P < 0.01). Taken altogether, proper dietary boron treatment might stimulate ostrich chick spleen development by promoting the Hsp70 expression level and inhibiting apoptosis, while a high amount of boron supplementation would impair the ostrich spleen structure by inhibiting Hsp70 expression level and promoting cell apoptosis.

Left jackknife position: a novel position for laparoscopic hepatectomy.

BACKGROUND: Laparoscopic hepatectomy for hepatocellular carcinoma (HCC) located in segment VI, VII, or VIII of the liver is usually difficult because of poor operative exposure, due to the unique anatomical structure. In this study, we evaluated the practice of laparoscopic hepatectomy with the left jackknife position for patients with HCC located in segment VI, VII, or VIII. METHODS: A total of 10 patients were enrolled to undergo laparoscopic hepatectomy with the left jackknife position. Tumors located in segment VI, VII, or VIII were assessed by preoperative dynamic computed tomography or magnetic resonance imaging. Operation time, intraoperative blood loss, postoperative fasting time, postoperative drainage time, major postoperative complications, and duration of postoperative hospital stay were recorded. RESULTS: All surgeries were successfully completed. None of the patients required conversion to open surgery during the procedure, and no serious postoperative complications were observed. The median tumor size was 31 mm (range 23-41 mm) in diameter, the mean operation time was 166 ± 38 min, the mean intraoperative blood loss was 220 ± 135 mL, and the median postoperative hospital stay was 4 days (range 2-7 days). CONCLUSIONS: For HCC located in segment VI, VII, or VIII, laparoscopic hepatectomy with this novel position-the left jackknife position-is safe and effective during tumor resection by exposing a sufficient operating field. Trial registration ClinicalTrials.gov ID: NCT02809287.

Prospective, single-center cohort study analyzing the efficacy of complete laparoscopic resection on recurrent hepatocellular carcinoma.

BACKGROUND: Laparoscopic hepatectomy is increasingly being used to treat hepatocellular carcinoma (HCC). However, few studies have examined the treatment of recurrent HCC in patients who received a prior hepatectomy. The present prospective study compared the clinical efficacy of laparoscopic surgery with conventional open surgery in HCC patients with postoperative tumor recurrence. METHODS: We conducted a prospective study of 64 patients, all of whom had undergone open surgery once before, who were diagnosed with recurrent HCC between June 2014 and November 2014. The laparoscopic group (n = 31) underwent laparoscopic hepatectomy, and the control group (n = 33) underwent conventional open surgery. Operation time, intraoperative blood loss, surgical margins, postoperative pain scores, postoperative time until the patient could walk, anal exsufflation time, length of hospital stay, and inpatient costs were compared between the two groups. The patients were followed up for 1 year after surgery, and relapse-free survival was compared between the two groups. RESULTS: All surgeries were successfully completed. No conversion to open surgery occurred in the laparoscopic group, and no serious postoperative complications occurred in either group. No significant difference in inpatient costs was found between the laparoscopic group and the control group (P = 0.079), but significant differences between the two groups were observed for operation time (116.7 ± 37.5 vs. 148.2 ± 46.7 min, P = 0.031), intraoperative blood loss (117.5 ± 35.5 vs. 265.9 ± 70.3 mL, P = 0.012), postoperative time until the patient could walk (1.6 ± 0.6 vs. 2.2 ± 0.8 days, P < 0.05), anal exsufflation time (2.1 ± 0.3 vs. 2.8 ± 0.7 days, P = 0.041), visual analogue scale pain score (P < 0.05), postoperative hepatic function (P < 0.05), and length of hospital stay (4.5 ± 1.3 vs. 6.0 ± 1.2 days, P = 0.014). During the 1-year postoperative follow-up period, 6 patients in each group had recurrent HCC on the side of the initial operation, but no significant difference between groups was observed in the recurrence rate or relapse-free survival. In the laparoscopic group, operation time, postoperative time until the patient could walk, anal exsufflation time, and inpatient costs were not different (P > 0.05) between the patients with contralateral HCC recurrence (n = 18) and those with ipsilateral HCC recurrence (n = 13). However, intraoperative blood loss was significantly less (97.7 ± 14.0 vs. 186.3 ± 125.6 mL, P = 0.012) and the hospital stay was significantly shorter (4.2 ± 0.7 vs. 6.1 ± 1.7 days, P = 0.021) for the patients with contralateral recurrence than for those with ipsilateral recurrence. CONCLUSIONS: For the patients who previously underwent conventional open surgical resection of HCC, complete laparoscopic resection was safe and effective for recurrent HCC and resulted in a shorter operation time, less intraoperative blood loss, and a faster postoperative recovery than conventional open surgery. Laparoscopic resection was especially advantageous for the patients with contralateral HCC recurrence.

Effect of Boric Acid Supplementation on the Expression of BDNF in African Ostrich Chick Brain.

The degree of brain development can be expressed by the levels of brain brain-derived neurotrophic factor (BDNF). BDNF plays an irreplaceable role in the process of neuronal development, protection, and restoration. The aim of the present study was to evaluate the effects of boric acid supplementation in water on the ostrich chick neuronal development. One-day-old healthy animals were supplemented with boron in drinking water at various concentrations, and the potential effects of boric acid on brain development were tested by a series of experiments. The histological changes in brain were observed by hematoxylin and eosin (HE) staining and Nissl staining. Expression of BDNF was analyzed by immunohistochemistry, quantitative real-time PCR (QRT-PCR), and enzyme linked immunosorbent assay (ELISA). Apoptosis was evaluated with Dutp-biotin nick end labeling (TUNEL) reaction, and caspase-3 was detected with QRT-PCR. The results were as follows: (1) under the light microscope, the neuron structure was well developed with abundance of neurites and intact cell morphology when animals were fed with less than 160 mg/L of boric acid (groups II, III, IV). Adversely, when boric acid doses were higher than 320 mg/L(groups V, VI), the high-dose boric acid neuron structure was damaged with less neurites, particularly at 640 mg/L; (2) the quantity of BDNF expression in groups II, III, and IV was increased while it was decreased in groups V and VI when compared with that in group I; (3) TUNEL reaction and the caspase-3 mRNA level showed that the amount of cell apoptosis in group II, group III, and group IV were decreased, but increased in group V and group VI significantly. These results indicated that appropriate supplementation of boric acid, especially at 160 mg/L, could promote ostrich chicks' brain development by promoting the BDNF expression and reducing cell apoptosis. Conversely, high dose of boric acid particularly in 640 mg/L would damage the neuron structure of ostrich chick brain by inhibiting the BDNF expression and increasing cell apoptosis. Taken together, the 160 mg/L boric acid supplementation may be the optimal dose for the brain development of ostrich chicks.

Increased Thymic Cell Turnover under Boron Stress May Bypass TLR3/4 Pathway in African Ostrich.

Previous studies revealed that thymus is a targeted immune organ in malnutrition, and high-boron stress is harmful for immune organs. African ostrich is the living fossil of ancient birds and the food animals in modern life. There is no report about the effect of boron intake on thymus of ostrich. The purpose of present study was to evaluate the effect of excessive boron stress on ostrich thymus and the potential role of TLR3/4 signals in this process. Histological analysis demonstrated that long-term boron stress (640 mg/L for 90 days) did not disrupt ostrich thymic structure during postnatal development. However, the numbers of apoptotic cells showed an increased tendency, and the expression of autophagy and proliferation markers increased significantly in ostrich thymus after boron treatment. Next, we examined the expression of TLR3 and TLR4 with their downstream molecular in thymus under boron stress. Since ostrich genome was not available when we started the research, we first cloned ostrich TLR3 TLR4 cDNA from thymus. Ostrich TLR4 was close to white-throated Tinamou. Whole avian TLR4 codons were under purify selection during evolution, whereas 80 codons were under positive selection. TLR3 and TLR4 were expressed in ostrich thymus and bursa of fabricius as was revealed by quantitative real-time PCR (qRT-PCR). TLR4 expression increased with age but significantly decreased after boron treatment, whereas TLR3 expression showed the similar tendency. Their downstream molecular factors (IRF1, JNK, ERK, p38, IL-6 and IFN) did not change significantly in thymus, except that p100 was significantly increased under boron stress when analyzed by qRT-PCR or western blot. Taken together, these results suggest that ostrich thymus developed resistance against long-term excessive boron stress, possibly by accelerating intrathymic cell death and proliferation, which may bypass the TLR3/4 pathway. In addition, attenuated TLRs activity may explain the reduced inflammatory response to pathogens under boron stress.

[Differential liver histopathological features of chronic HBV infection patients with normal and mildly elevated serum ALT].

To study the liver histopathological features that are distinctive between chronic hepatitis B virus (HBV) infection patients who have normal serum alanine aminotransferase (ALT)/asparatate aminotransferase (AST) and those with mildly elevated serum ALT/AST. One-hundred-and-thrity-four chronic HBV infection patients with normal serum ALT/AST and 165 chronic HBV infection patients with mildly elevated serum ALT/AST were included in the study. Liver biopsies were performed and used to assess the histological changes by hematoxylin-eosin and reticular fiber staining; mild to severe scoring for inflammation was made as grade G0-G4 and for fibrosis stage as S0-S4. HBV DNA levels were detected by fluorescent quantitative PCR. HBV serological markers were examined by chemiluminescence. The mildly elevated serum ALT/AST group had more male patients than the normal serum ALT/AST group. In the normal serum ALT/AST group, 50.0% (67/134) of the patients had moderate histological changes and only 3.0% (4/134) had severe changes (G3-4 and/or S3-4). In the mildly elevated ALT/AST group, 65.7% (174/265) of patients had moderate histological changes and 16.2% (43/265) had severe changes (G3-4 and/or S3-4). Hepatic inflammation and fibrosis were significantly more severe in the mildly elevated serum ALT/AST group than in the normal ALT/AST group (x2 = 26.386, P less than 0.01; x2 = 15.299, P less than 0.01). In the normal ALT/AST group, the severity of inflammation and fibrosis were positively correlated with age (rs = 0.620, P less than 0.01; rs = 0.347, P less than 0.01). In the mildly elevated ALT/AST group, the severity of inflammation and fibrosis were negatively correlated with age (rs = -0.807, P less than 0.01; rs = -0.557, P less than 0.01). In both groups, the severity of inflammation and fibrosis were negatively correlated with HBV DNA levels (rs = -0.215, P less than 0.01, rs = -0.527, P less than 0.01, rs = -0.951, P less than 0.01; rs = -0.715, P less than 0.01) and were not positively correlated with HBeAg. The majority of the chronic HBV infection patients with normal serum ALT/AST and those with mildly elevated serum ALT/AST had moderate liver pathological changes. All patients with low HBV DNA levels were closely followed-up, regardless of HBeAg-positive status.

[Study on the copy numbers and mRNA expression levels of the programmed death-1 gene in chronic hepatitis B patients].

OBJECTIVE: To study the copy numbers and mRNA expression levels of the Programmed Death-1 gene in chronic hepatitis B patients and to analyze the differences of the copy numbers and mRNA expression levels of the gene in patients with different clinical outcomes. METHODS: Real time PCR was adopted to detect the PD-1 gene copy numbers and their mRNA expressions in peripheral blood mononuclear cells (PBMCs) from 27 samples from healthy donors in Control group, 31 samples from chronic asymptomatic HBV carriers (ASC, n=31), 19 samples from chronic severe hepatitis B patients (CSH, n=19) and 29 samples from Primary hepatitis B Virus-related hepatocarcinoma (PHC, n=29). The differences and relationship of copy numbers and their mRNA expression levels among those groups were compared and analyzed by adopting Chi-square test and Rank sum test. RESULTS: PD-1 gene copy number deviated from 0 copy to 3 copies among all the 106 samples. In control group, ASC group, CSH group and PHC group, the percentages of cases of haploid (single) were 37.0%, 35.5%, 26.3% and 6.9%, respectively, the percentages of cases of diploid (double) were 55.5%, 58.0%, 63.2% and 82.8%, respectively, and the percentages of cases of triploid (triple) were 3.7%, 6.5%, 10.5% and 10.3%, respectively. The percentage of cases of polyploid (diploid and triploid) in control group, ASC group, CSH group and PHC group were 59.3%, 64.5%, 73.7% and 93.1%, respectively. The different distribution of PD-1 gene copy number of polyploid was significant in total samples (x2=9.583, P<0.05). Compared with Control Group and ASC group, the percentage of cases of polyploid in PHC group was lower with the x2 equals to 8.985 and 7.215 respectively and both with P less than 0.05. The difference between the two groups was statistically significant. The mean PD-1 gene copy numbers for these four groups were 1.59+/-0.63, 1.70+/-0.52, 1.84+/-0.60 and 2.00+/-0.37 while the median were 0.002 54, 0.002 72, 0.002 55 and 0.001 33 respectively. Except the control group, there was a uptrend in the other three groups while PD-1 gene mRNA expression presented a downtrend. The mean of PD-1 gene copy numbers of 2 and their mRNA expression levels were 19.59, 32.57 and 33.22 for PHC, CSH and ASC groups among which PHC group had the lowest value, there was significant differences found in the comparison with F=5.395 and P<0.05. CONCLUSION: PD-1 gene copy numbers and their mRNA expression levels were different in chronic HBV infected patients with different transformation. It is valuable to follow up the patients with more than 1 copy number of PD-1 gene in long term.