[Determination of 8 N-nitrosamines in the workplace air by GC-MS/MS method].

Objective: To establish a method for the determination of eight N-nitrosamines (N-nitrosodimethylamine, N-nitrosodimethylamine, N-nitrosomethylmethylamine, N-nitrosodibutylamine, N-nitrosopropylamine, N-nitrosomorpholine, N-nitrosodianiline and N-nitrosopiperidine) in the air of workplace by gas chromatography-tandem mass spectrometry (GC-MS/MS) . Methods: From January to August 2023, eight N-nitrosamines in the air of workplace were collected by ThermoSorb/N column, eluted with 4 ml methanol-dichloromethane (1∶1 volume ratio), separated by VF-624 ms capillary column, detected by multiple reaction monitoring mode and quantified by external standard method. The detection limit and precision of the method were also analyzed. Results: The linear range of the method for the determination of eight N-nitrosamines was 1.0-20.0 µg/L, the correlation coefficient was 0.9993-0.9999, the detection limit was 0.051-0.132 µg/L, and the minimum quantitative concentration was 0.030-0.078 µg/m(3) (calculated by collecting 22.5 L of air sample and eluting with 4.0 ml stripping liquid). The within-run precisions were 2.05%-6.89% and the between-run precisions were 2.41%-8.26%. The desorption rates were 67.20%-102.60%. The sample can be kept at least 7 days at 4 ℃. Conclusion: GC-MS/MS method for the determination of eight N-nitrosamines in workplace air has high sensitivity and good precision, and can accurately determine the content of eight N-nitrosamines in workplace air.

Circular RNA circRNA_0000285 promotes cervical cancer development by regulating FUS.

OBJECTIVE: Recently, the vital role of circular RNAs (circRNAs) in human diseases has attracted much attention. The aim of this research was to verify the potential role of circRNA_0000285 in the development of cervical cancer (CC). PATIENTS AND METHODS: CircRNA_0000285 expression in both CC cells and tissue samples was detected by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). Functional experiments were performed, including cell counting kit-8 (CCK-8) assay, cell cycle assay and transwell assay. Meanwhile, the underlying mechanism was explored through qRT-PCR and Western blot assay, respectively. In addition, the function of circRNA_0000285 was identified in vivo. RESULTS: CircRNA_0000285 expression level was significantly higher in CC samples than that of corresponding normal tissues. Moreover, the growth and migration abilities of CC cells were significantly inhibited after circRNA_0000285 was knocked down in vitro. Furthermore, the expression of FUS was remarkably down-regulated after knockdown of circRNA_0000285. Subsequent results indicated that the expression level of FUS was positively correlated with the expression of circRNA_0000285 in CC tissues. In addition, the knockdown of circRNA_0000285 significantly inhibited the formation and metastasis of CC in nude mice. CONCLUSIONS: CircRNA_0000285 could enhance the proliferation and metastasis of CC by up-regulating FUS, which might be a potential therapeutic target for CC treatment.

[The role of aquaporin 4 antibody in the injury of retinal microstructure in neuromyelitis optica spectrum disorders].

Objective: To evaluate the injury of retinal microstructure using optical coherence tomography (OCT) and investigate the role of aquaporin 4 antibody (AQP4 Ab) in this injury process. Methods: Forty patients with neuromyelitis optica spectrum disorders (NMOSD) were retrospectively studied, each of whom reported at least one episode of optic neuritis (ON), namely 59 ON eyes involved in all. All patients were divided into two subgroups based on AQP4 Ab tests including 25 patients (37 ON eyes) with AQP4 positive (Ab(+)/NMOSD group) and 15 patients (22 ON eyes) negative (Ab(-)/NMOSD group). In addition, 10 healthy controls (20 eyes) matched for age and sex (HC group) were analyzed. Spectral domain optical coherence tomography (SD-OCT) was used to quantify peripapillary retinal nerve fiber layer (RNFL). Nonparametric test was used to compare differences between groups. Results: Age distribution and gender ratio were comparable in three groups (P>0.05). Visual acuity in ON eyes of Ab(+)/NMOSD group was worse than that of Ab(-)/NMOSD group (P=0.02). There were no significant differences between Ab(+)/NMOSD and Ab(-)/NMOSD in aspects of disease duration (2.6 vs. 1.9 year), ON episodes (2 vs. 1), longitudinal extensive transverse myelitis (LETM) ratio (48.0% vs. 66.7%), NMOSD specific intracranial lesions ratio (32.0% vs. 53.3%), positive autoimmune antibody ratio (52.0% vs. 20.0%) (P= 0.13, 0.08, 0.25, 0.18, 0.06, respectively). The thickness of temporal, superior, nasal, inferior and average RNFL in ON eyes of both Ab(+)/NMOSD and Ab(-)/NMOSD group were thinner than those in eyes of HC group (all P<0.05). The thickness of superior and inferior RNFL in ON eyes of Ab(+)/NMOSD were 61.0 µm and 62.0 µm, which was thinner than those of Ab(-)/NMOSD 94.5 µm and 97.0 µm (P=0.03 and 0.01, respectively). Conclusions: RNFL reflects the injury of retinal microstructure in NMOSD patients. AQP4 Ab seropositivity is correlated to the severity of RNFL damage, implying the potential role of AQP4 Ab in this pathological process.

[Clinical features of late-onset neuromyelitis optica spectrum disorders].

Objective: To study the clinical features of late-onset neuromyelitis optica spectrum disorders(LONMOSD). Methods: Twenty-eight patients with LONMOSD and fifty-one patients with early-onset neuromyelitis optica spectrum disorders(EONMOSD) hospitalized in Navy General Hospital from January 2014 to May 2017 were enrolled and were followed up by telephone or outpatient visiting. The clinical manifestations, laboratory examinations and imaging features of the two groups were compared. Results: (1)The average age of onset in group LONMOSD was(59±6) years old, and 24 cases were female(85.7%). (2) The history of prodromal infection in LONMOSD patients was less reported than that in group EONMOSD(14.3 vs 37.3%, P<0.05), but concomitant diseases were more common in LONMOSD patients(53.6% vs 3.9%, P<0.05). (3) In group of LONMOSD, the patients with transverse myelitis(TM )as the first symptom were less than that of EONMOSD group (39.3% vs 64.7%, P<0.05). (4) There was no significant difference in EDSS score either in acute or remission stage, laboratory and imaging findings between the two groups. Conclusion: Patients with LONMOSD have less history of prodromal infection, and those with TM as the first symptom are less than EONMOSD patients.

[The clinical analyses of neuromyelitis optica spectrum disorder initially presenting with area postrema syndrome in 14 patients].

Objective: To explore the clinical characteristics of the neuromyelitis optica spectrum disorders (NMOSD) with the area postrema syndrome as the initial symptom. Methods: A total of 14 cases were enrolled in the study with the diagnose of NMOSD and the area postrema syndrome as the initial symptom. All the clinical data and imaging profiles by the contrasted magnetic resonance imaging (MRI) of the head and spinal cord were collected and analyzed. Results: The median age of onset was (38.1±17.0) years old and the gender ratio of female to male was 10∶4. The serum aquaporin-4(AQP4)-IgG was positive in 11 subjects and several autoimmune antibodies was positive in 7 subjects. The lesions revealed by MRI of the head mainly located in the area postrema and ependymal periphery which often presented as the linear medullary lesion, while linear lesions over three pieces of vertebra were shown by MRI of the spinal cord which mainly in the grey matter and with a"H" shape around the spinal central canal. Misdiagnose happened in 11 subjects with seven of gastroesophageal reflux disease, two of neurogenic vomiting, one of spinal cord tuberculosis and one of stroke. Conclusions: NMOSD should be considered in patients with unexplained intractable nausea, vomiting and/or hiccups lasted for 48 hours or above, especially in those with positive nervous signs. Contrasted MRI and serum AQP4-IgG need to be performed in the suspected patients. Early detection is crucial for patients with NMOSD.

[Analysis of the clinical features of vestibular migraine with MRI changes].

Objective: To investigate the clinical features of vestibular migraine with MRI changes. Methods: The clinical data of 150 patients with vestibular migraine completing MRI examination in navy general hospital from August 2008 to August 2016 were retrospectively analyzed.Clinical features of 94 (62.7%) vestibular migraine patients with MRI changes were collected and analyzed according to the medical history, clinical symptoms and signs, as well as the result of auxiliary examinations. Results: The manifestations of MRI in vestibular migraine patients were mainly multiple punctate equal T(1) and long T(2) signals or high signals on FLAIR (fluid attenuated inversion recovery) in subcortical white matter. The average age of the patients at the visit was (52±15) years old, with the male to female ratio of 1∶3.1. The occurrence of vertigo and migraine were in no particular order, with 54.3% patients presenting migraine several years before vertigo. The duration of vertigo was mostly 24 hours to 72 hours (31.9%). The visual aura (55.3%), photophobia and phonophobia (67.0%) were the most frequently associated symptoms. Conclusions: The vestibular migraine patients with MRI changes have its own specific clinical characteristics and are mainly seen in female with visual aura. The associated symptoms of vestibular migraine appear mostly during the process of vertigo. The pathogenesis of cerebral white matter hyperintensities (WMHs) is not clear.

Osteogenesis induced in goat bone marrow progenitor cells by recombinant adenovirus coexpressing bone morphogenetic protein 2 and basic fibroblast growth factor.

Bone morphogenetic protein 2 (BMP2) and basic fibroblast growth factor (bFGF) have been shown to exhibit a synergistic effect to promote bone repair and healing. In this study, we constructed a novel adenovirus with high coexpression of BMP2 and bFGF and evaluated its effect on osteogenic differentiation of goat bone marrow progenitor cells (BMPCs). Recombinant adenovirus Ad-BMP2-bFGF was constructed by using the T2A sequence. BMPCs were isolated from goats by density gradient centrifugation and adherent cell culture, and were then infected with Ad-BMP2-bFGF or Ad-BMP2. Expression of BMP2 and bFGF was detected by ELISA, and alkaline phosphatase (ALP) activity was detected by an ALP assay kit. In addition, von Kossa staining and immunocytochemical staining of collagen II were performed on BMPCs 21 days after infection. There was a high coexpression of BMP2 and bFGF in BMPCs infected with Ad-BMP2-bFGF. Twenty-one days after infection, ALP activity was significantly higher in BMPCs infected with Ad-BMP2-bFGF than in those infected with Ad-BMP2. Larger and more mineralized calcium nodules, as well as stronger collagen II staining, were observed in BMPCs infected with Ad-BMP2-bFGF than in those infected with Ad-BMP2. In summary, we developed a novel adenovirus vector Ad-BMP2-bFGF for simultaneous high coexpression of BMP2 and bFGF, which could induce BMPCs to differentiate efficiently into osteoblasts.

Osteogenesis induced in goat bone marrow progenitor cells by recombinant adenovirus coexpressing bone morphogenetic protein 2 and basic fibroblast growth factor

Bone morphogenetic protein 2 (BMP2) and basic fibroblast growth factor (bFGF) have been shown to exhibit a synergistic effect to promote bone repair and healing. In this study, we constructed a novel adenovirus with high coexpression of BMP2 and bFGF and evaluated its effect on osteogenic differentiation of goat bone marrow progenitor cells (BMPCs). Recombinant adenovirus Ad-BMP2-bFGF was constructed by using the T2A sequence. BMPCs were isolated from goats by density gradient centrifugation and adherent cell culture, and were then infected with Ad-BMP2-bFGF or Ad-BMP2. Expression of BMP2 and bFGF was detected by ELISA, and alkaline phosphatase (ALP) activity was detected by an ALP assay kit. In addition, von Kossa staining and immunocytochemical staining of collagen II were performed on BMPCs 21 days after infection. There was a high coexpression of BMP2 and bFGF in BMPCs infected with Ad-BMP2-bFGF. Twenty-one days after infection, ALP activity was significantly higher in BMPCs infected with Ad-BMP2-bFGF than in those infected with Ad-BMP2. Larger and more mineralized calcium nodules, as well as stronger collagen II staining, were observed in BMPCs infected with Ad-BMP2-bFGF than in those infected with Ad-BMP2. In summary, we developed a novel adenovirus vector Ad-BMP2-bFGF for simultaneous high coexpression of BMP2 and bFGF, which could induce BMPCs to differentiate efficiently into osteoblasts.

Value of a skin island flap as a postoperative predictor of vascularized fibula graft viability in extensive diaphyseal bone defect reconstruction.

PURPOSE: To evaluate the feasibility and reliability of free vascularized fibular graft with skin island flap for reconstruction of large diaphyseal bone defect. METHOD: The clinical results of vascularized fibular graft and experiences related to the importance and reliability of a monitoring island flap for the reconstruction of various long-bone defects were reviewed in 87 patients. RESULTS: Bony reconstruction was achieved in 82 of the 87 patients. Arterial thrombosis of anastomosed vessel in two patients and venous congestion of monitoring flap in nine patients occurred in the early postoperative periods. All of them were managed by immediate thrombectomy and reanastomosis, alternatively the thrombotic veins were replaced by new veins to anastomose with the superficial veins in five patients. Partial flap necrosis was noted in six patients, but additional surgical intervention was not required. The vascularized fibula survived and bony fusion was achieved in all patients. Postoperative stress fractures of the fibula graft occurred in 19 (21.8%) patients (once in seven patients, twice in five patients, three or more times in seven) as the mechanical stress to the graft increased. Included fracture on the tibia in 12 patients, humerus in one and femur in six. Treatments included casting in 11 patients, percutaneous pinning in one case, and adjustment of external fixator in seven patients. Bony union was finally achieved an average of 9.6 months after fracture. CONCLUSIONS: Correct alignment between the recipient bone and the external fixator is a prerequisite to preventing graft fracture. Vascularized fibula transfer is a valuable procedure for long-bone defects, and a skin island-monitoring flap is a simple, extremely useful, and reliable method for assessing the vascular status of vascularized fibula. LEVEL OF EVIDENCE: Level IV. Retrospective study.

Manipulation of monoubiquitin improves salt tolerance in transgenic tobacco.

Ubiquitin (Ub) is regarded as a stress protein involved in many stress responses. In this paper, sense and antisense transgenic tobacco plants, as well as the wild type and vector control, were used to study the role of Ub in salt tolerance of plants. In sense Ta-Ub2 transgenic tobacco plants, there was higher expression of Ub protein conjugates than in the wild type and vector control, but the reverse trend was observed in antisense Nt-Ub1 transgenic plants. The germination rate of tobacco seed, growth status and photosynthesis of the tobacco plants suggested that over-expressing Ub promoted the growth of transgenic tobacco plants and enhanced their salt tolerance, but the opposite effect was seen in plants with repressed Ub expression. Changes in antioxidant capacity may be one of the mechanisms underlying Ub-regulated salt tolerance. Furthermore, improved tolerance to a combination of stresses was also observed in the sense transgenic tobacco plants. These findings imply that Ub is involved in the tolerance of plants to abiotic stress.