Objective To prepare a variety of quality control(QC)materials for SARS-CoV-2 variants as an addition to the conven-tional SARS-CoV-2 nucleic acid QC products for the laboratory detection of mutant strains by optimizing the preparation and purification process of MS2 phage virus-like particle(VLP)technique,and evaluate their performances.Methods The typical mutation sequence fragments or full length S genes were designed and synthesized according to the genomic information of SARS-CoV-2 variants.Then,they were inserted into the downstream of maturase gene,coat protein and the pac-site of MS2 phage to construct a series of recombi-nant expression vectors.After induced by the prokaryotic expression system,the VLP products were purified through the polyethylenei-mine precipitation,ultrafiltration,nuclease digestion,and gel filtration chromatography.The obtained VLP were validated by the nucle-ic acid electrophoresis,protein electrophoresis,protein concentration determination,and fluorescence PCR,and their performances such as nucleic acid residue and stability were also evaluated.Results A total of 10 kinds of VLP containing the targeted sequences of the gene to be tested were prepared.The length of the foreign sequence wrapped in them ranged from 297 bp to 3 822 bp,which could be combined into a variety of QC materials for the mutation detection of different SARS-CoV-2 variants.The prepared VLP QC materials could not be effectively amplified without nucleic acid extraction or reverse transcription steps during the routine nucleic acid detection.The simulated QC samples remained stable after repeated freeze-thaw cycles.They could be stored stably for 2 months at 25 ℃ and 4 weeks at 37 ℃.Conclusion The established preparation and combined purification process of VLP QC materials can encapsulate vari-ous exogenous nucleic acid sequences with different lengths into the viral coat protein to form VLP,with high production efficiency.The VLP QC products prepared by the above process have stable performance and almost no residual exogenous nucleic acid,which can ef-fectively meet clinical requirements and ensure the quality of laboratory testing.

Objective To prepare a variety of quality control(QC)materials for SARS-CoV-2 variants as an addition to the conven-tional SARS-CoV-2 nucleic acid QC products for the laboratory detection of mutant strains by optimizing the preparation and purification process of MS2 phage virus-like particle(VLP)technique,and evaluate their performances.Methods The typical mutation sequence fragments or full length S genes were designed and synthesized according to the genomic information of SARS-CoV-2 variants.Then,they were inserted into the downstream of maturase gene,coat protein and the pac-site of MS2 phage to construct a series of recombi-nant expression vectors.After induced by the prokaryotic expression system,the VLP products were purified through the polyethylenei-mine precipitation,ultrafiltration,nuclease digestion,and gel filtration chromatography.The obtained VLP were validated by the nucle-ic acid electrophoresis,protein electrophoresis,protein concentration determination,and fluorescence PCR,and their performances such as nucleic acid residue and stability were also evaluated.Results A total of 10 kinds of VLP containing the targeted sequences of the gene to be tested were prepared.The length of the foreign sequence wrapped in them ranged from 297 bp to 3 822 bp,which could be combined into a variety of QC materials for the mutation detection of different SARS-CoV-2 variants.The prepared VLP QC materials could not be effectively amplified without nucleic acid extraction or reverse transcription steps during the routine nucleic acid detection.The simulated QC samples remained stable after repeated freeze-thaw cycles.They could be stored stably for 2 months at 25 ℃ and 4 weeks at 37 ℃.Conclusion The established preparation and combined purification process of VLP QC materials can encapsulate vari-ous exogenous nucleic acid sequences with different lengths into the viral coat protein to form VLP,with high production efficiency.The VLP QC products prepared by the above process have stable performance and almost no residual exogenous nucleic acid,which can ef-fectively meet clinical requirements and ensure the quality of laboratory testing.

Purpose To investigate the relationship between imaging characteristics and shunt grading in acute cerebral infarction patients with right-to-left shunt.Materials and Methods The clinical and imaging data of 697 acute cerebral infarction patients who were hospitalized in the Second Affiliated Hospital of Nanchang University from January 2019 to December 2021 were collected,and the patients were divided into right-to-left shunt negative group(443 cases)and right-to-left shunt positive group(254 cases)according bubble test,compared the difference in demographic data and imaging characteristics between the two groups.In the shunt positive group,the imaging characteristics of the two groups of patients and their relationship with the shunt grade were analyzed.Results There were no statistical differences in gender,age,smoking,hyperlipidemia and history of coronary heart disease between the two groups(P>0.05),but there were statistical differences in the history of hypertension(OR=0.533,95%CI 0.380-0.748),diabetes(OR=0.649,95%CI 0.422-0.999)and cerebral infarction(OR=0.275,95%CI 0.136-0.555)(all P<0.05).The imaging characteristics of the two groups of patients showed that there were significant differences in the number of infarct foci(χ2=53.957,4.219;both P<0.05),but there was no statistical difference in the infarct location between the two groups(P>0.05).Among the patients with acute cerebral infarction combined with positive bubble test,the imaging manifestations of grading I and II patients were mainly unilateral anterior circulation or posterior circulation infarction,and deep and single infarction were more common.In patients with grading III and IV shunts,multiple infarcts with combined bilateral anterior and posterior circulations were more common.Conclusion The imaging manifestations of patients with acute cerebral infarction combined with right-to-left shunt have their own characteristics.Bubble test has high sensitivity in the diagnosis of right-to-left shunt.The comprehensive analysis can provide certain guidance for clinical practice.

Spermatogenesis is a continual process that occurs in the testes, in which diploid spermatogonial stem cells (SSCs) differentiate and generate haploid spermatozoa. This highly efficient and intricate process is orchestrated at multiple levels. N⁶-methyladenosine (m⁶A), an epigenetic modification prevalent in mRNAs, is implicated in the transcriptional regulation during spermatogenesis. However, the dynamics of m⁶A modification in non-rodent mammalian species remains unclear. Here, we systematically investigated the profile and role of m⁶A during spermatogenesis in pigs. By analyzing the transcriptomic distribution of m⁶A in spermatogonia, spermatocytes, and round spermatids, we identified a globally conserved m⁶A pattern between porcine and murine genes with spermatogenic function. We found that m⁶A was enriched in a group of genes that specifically encode the metabolic enzymes and regulators. In addition, transcriptomes in porcine male germ cells could be subjected to the m⁶A modification. Our data show that m⁶A plays the regulatory roles during spermatogenesis in pigs, which is similar to that in mice. Illustrations of this point are three genes (SETDB1, FOXO1, and FOXO3) that are crucial to the determination of the fate of SSCs. To the best of our knowledge, this study for the first time uncovers the expression profile and role of m⁶A during spermatogenesis in large animals and provides insights into the intricate transcriptional regulation underlying the lifelong male fertility in non-rodent mammalian species.
Animals ; Male ; Mice ; Cell Differentiation/genetics* ; Mammals/metabolism* ; Methylation ; RNA, Messenger/metabolism* ; Spermatogenesis/genetics* ; Spermatozoa/metabolism* ; Swine/genetics* ; Testis/metabolism* ; Transcriptome ; RNA Methylation/genetics*

OBJECTIVE：To observe the ef fectiveness and safety of bosentan in the treatment of hypoxic pulmonary hypertension（HPH）in neonates. METHODS ：From Jan. 2014 to Mar. 2019，a total of 82 HPH neonates hospitalized in the department of neonatology of our hospital were selected as research subjects. According to whether or not receiving bosentan therapy，50 cases were included into bosentan group and 32 cases into non-bosentan group. Meanwhile ，another 25 non-HPH neonates with serum sample retention time and general information such as gestational age at birth and day age matching the HPH group were selected as the control group. All neonates with HPH were given continuous intravenous infusion of Dopamine hydrochloride injection 5 mg/（kg·min）until PASP was normal. On this basis, neonates in the bosentan group were additionally given Bosentan tablets 1 mg/kg（fed after dissolving with appropriate amount of water for injection ）for q 12 h，72 h. The relationship between serum ET- 1 levels of neonates with HPH and PASP was analyzed ，as well as PASP before and after treatment and therapeutic efficacy between bosentan and non-bosentan groups ，the changes of arterial blood gas indexes and ADR in 3 groups were compared. RESULTS ：Before treatment ，the serum ET- 1 levels of bosentan group was （164.3±115.3）pg/mL，which was significantly higher than （41.9±3.7）pg/mL of control group and positively correlated with PASP level （r＝0.864，P＜0.001）. Total response rate of bosentan group was 90.00％，which was significantly higher than 71.88％ of non-bosentan group （P＜0.05）. After 72 h of treatment ，PASP of 2 groups was decreased significantly ，compared with before treatment （P＜0.001），and the bosentan group was significantly lower than the non-bosentan group （P＜0.05）. The PaO 2，SaO2，PaCO2 and OI in 3 groups was significantly improved compared with that before treatment （P＜0.001），and the PaO 2，SaO2 and OI in the bosentan group was significantly higher than that in the non-bosentan group （P＜0.05）. During the treatment period of bosentan and within one week after drug withdrawal，there was no significant change in serum LDH ，AST，ALT and Scr levels in neonates. There was no statistically significant difference in the incidence of feeding intolerance ，anemia，reduced WBC and reduced PLT in 3 groups（P＞0.05）. CONCLUSIONS： Bosentan can improve the oxygenation status of neonates with HPH, reduce PA SP，and short-termmedication is safe. com

Spinal cord injuries is an extremely serious central nervous system injury. The clinical prognosis is very poor. Patients are often associated with lifelong disabilities or paralysis. Regulating NSCs to repair spinal cord injuries is unanimously considered to be a very potential option for the treatment of this type of disease. China has a large population and a large number of patients with spinal cord injuries. Actively regulating spinal cord NSCs is of great significance for the regeneration and repair of spinal cord injuries. Endogenous NSCs avoid many disadvantages of exogenous stem cell transplantation, and have a broader prospect in the treatment of spinal cord injuries. The Wnt signaling pathway plays a very important role in the differentiation of NSCs and the development of the nervous system. However, the molecular mechanism of the proliferation and differentiation of NSCs during regeneration and repair after spinal cord injuries is still not fully understood. This article mainly describes the research progress of Wnt pathway regulating NSCs in the regeneration and repair of spinal cord injuries.

Objective: Analysis of the effect of triggering receptor-1 expressed on myeloid cells (TREM-1) in nonalcoholic fatty liver disease (NAFLD) and the mechanism. Methods: The oleic acid-treated HepG2 cells were divided into model group, overexpression group, interference group A, interference group B and negative control group. The mouse model of NAFLD was generated and randomly divided into (nuclear factor-κB) NF-κB inhibition group, protein kinase B (AKT) inhibition group, knockout group A, knockout group B and control group. The expression of inflammatory factors and TREM-1 in liver tissue was detected by PCR, and fat accumulation was detected by oil red O staining. Western blotting was used to detect the expression of TREM-1 and signaling pathway proteins, and HE staining was used to detect liver tissue changes. Results: TREM-1 was up-regulated in liver tissue of NAFLD mice [(0.936±0.127) vs. (0.432±0.105)] and in oleic acid-treated HepG2 cells. In oleic acid-treated HepG2 cells, overexpression of TREM-1 increased inflammatory factor expression and increased lipid droplets; inhibition of TREM-1 expression decreased inflammatory factor expression, and lipid droplets decreased. Knockout of TREM-1 and inhibition of NF-κB in NAFLD mice reduced hepatocyte inflammatory factor expression and reduced liver damage; knockout of TREM-1 and inhibition of AKT reduced liver tissue lipids and drops accumulate. Conclusions The overexpression of TREM-1 in NAFLD mice liver tissue can regulate inflammatory factor expression and lipid droplets through NF-κB and AKT signal pathway. TREM-1 might be a potential therapeutic target of NAFLD.

To investigate the values of orbital single photon emission computed tomography/computerized tomography (SPECT/CT) with 99mTc-diethylene triamine pentaacetic acid (99mTc-DTPA) on the evaluation of lacrimal gland inflammation in patients with thyroid associated ophthalmopathy (TAO).
 Methods: A total of 58 TAO patients were retrospectively recruited for this study, all of whom were categorized into all active group and all inactive group based on the clinical active score (CAS). Another 12 patients with the negative images on SPECT/CT served as a normal control group (NC). All patients were undergone the 99mTc-DTPA orbit SPECT/CT. Quantitative parameters of lacrimal gland including width, length, volume, and the count ratio of region of interest (ROI) drawn on lacrimal gland to the region of occipital brain (target/non-target ratio, T/NT), were measured on axial and coronal slices, respectively. Quantitative parameters were compared among the 3 groups, and the diagnostic value on discrimination of TAO patients from inactive to active ones was evaluated.
 Results: All parameters in TAO patients (except the length on coronal slices and the mean radioactive counts of axial T/NT in the TAO inactive group) were much greater than those in the NC group (P<0.05). All parameters in the TAO active group (except the length on axial and coronal slices) were much greater than those in the TAO inactive group (P<0.05). There were significant lineal positive correlations between the parameters in all TAO patients (except the length on coronal slices ) and CAS. The best diagnostic value was detected by the max radioactive counts of axial T/NT (area under the curve=0.82, P<0.01).
 Conclusion: As an invasive imaging modality, 99mTc-DTPA orbital SPECT/CT is helpful to estimate the lacrimal gland inflammation and to assess the disease activity in TAO patients.
Graves Ophthalmopathy ; Humans ; Inflammation ; Lacrimal Apparatus ; Orbit ; Retrospective Studies ; Tomography, Emission-Computed, Single-Photon ; Tomography, X-Ray Computed

To compare diagnostic value of 4 kinds of target-to-nontarget ratio (T/NT), and to choose a better one to assess thyroid associated ophthalmopathy (TAO) activity.
 Methods: The clinical data were collected for 29 newly-diagnosed patients (58 eyes) with TAO who underwent orbital 99mTc-DTPA single photon emission computed tomography/computed tomography (SPECT/CT) fusion images according to the clinical activity score (CAS). They were divided into an active group (18 cases, 36 eyes), an inactive group (11 cases, 22 eyes), and a control group (9 cases, 18 eyes). Diagnostic value of orbital/occipital lobe radioactive uptake count ratio (T/NT1), orbital/occipital radioactive uptake count ratio (T/NT2), orbital/thalamus radioactive uptake count ratio (T/NT3), and orbital/cerebellar radioactivity uptake count ratio (T/NT4) were calculated, and the CAS of Spearman rank correlation and receiver operating characteristic (ROC) curve were analyzed.
 Results: T/NT1, T/NT2 and CAS were correlated (r1=0.873, r2=0.527; P<0.001), with the better correlation between T/NT1 and CAS and there was no correlation between T/NT3, T/NT4 and CAS (r3=0.039, r4=0.090; P>0.05). Area under the ROC curve of T/NT1 was 0.860, area under the ROC curve of T/NT2 was 0.581, and the accuracy for T/NT1 on TAO activity was good. T/NT1=9.74 could be used as active threshold for judge of TAO in patients.
 Conclusion: There is a good correlation between T/NT1 and CAS. TAO activity assessment possesses high diagnostic value, and SPECT/CT together with imaging anatomical location is more accurate.
Case-Control Studies ; Graves Ophthalmopathy ; diagnostic imaging ; Humans ; Orbit ; diagnostic imaging ; ROC Curve ; Radiopharmaceuticals ; Statistics, Nonparametric ; Technetium Tc 99m Pentetate ; Tomography, Emission-Computed, Single-Photon ; methods

Objective To analyze the changes of serum hypoxia-inducible factor-1α(HIF-1α),vas-cular endothelial growth factor ( VEGF) and endothelin-1 ( ET-1) levels in neonates with hypoxia-induced pulmonary hypertension(HPH),and to explore the roles of three factors in the pathogenesis of HPH in neo-nates. Methods A total of 50 neonates with HPH in the neonatal intensive care unit as HPH group and an-other 25 non-HPH hospitalized neonates with similar clinical data in the same period as control group were enrolled from January 2014 to December 2017. The levels of serum HIF-1α,VEGF and ET-1 were deter-mined by enzyme linked immunosorbent assay when neonates were diagnosed as HPH and their pulmonary artery systolic blood pressure(PASP) decreased to 35 mmHg below respectively. The changes of three factors levels were analyzed and compared with those of control group at the same time point. Results (1) The levels of serum HIF-1α,VEGF and ET-1 of HPH group were significantly higher than those of the control group (F value were 151. 97,43. 31,and 129. 56 respectively,all P<0. 01). Furthermore,the more serious the grade of HPH,the higher the levels of three factors. The differences were statistically significant (all P<0. 01). (2) After PASP of neonates in HPH group decreased to 35 mmHg or below,the levels of serum HIF-1α,VEGF and ET-1 also significantly decreased,and the differences were statistically significant ( all P <0. 01). However,there were no statistically significant differences compared with those of the control group for the levels of serum HIF-1α and VEGF(both P>0. 05) in addition to serum ET-1 levels (F＝14. 98,P<0. 05). Conclusion High levels of HIF-1α,VEGF and ET-1 caused by hypoxia may play an important role in neonatal HPH.