[IgG4-related ophthalmic disease: three-case report].

The first patient complained visual loss of left eye for 1 week. The second patient complained visual loss of right eye for 2 months with conjunctival edema for more than 10 days. The third patient complained redness and swelling of both eyes for 1 year, poor appetite and fatigue for 4 months. All three patients were male with binocular involvement. Enlargement of lacrimal glands in both eyes with bulbar conjunctival edema were observed in 2 patients and binocular eyelid swelling in 1 patient, all 3 patients were associated with external orbital tissue involvement, which were diagnosed as IgG4-related ophthalmic diseases. After treatment with glucocorticoid, the symptoms of the patients were improved, and the patients were followed up for 58, 54, and 60 months, respectively. Among the 3 patients, 2 had recurrence for once, and 1 had recurrence 3 for times. (Chin J Ophthalmol, 2021, 57: 375-378).

[miR-758-3p inhibits the proliferation and invasion of non-small cell lung cancer cells by targeting NUSAP1].

Objective: To investigate the biological function of miR-758-3p and the underlying mechanism in non-small cell lung cancer (NSCLC). Methods: miR-758-3p mimics was transfected to A549 NSCLC cells, miRNA control was used as a negative control, cells transfected with nucleolar and spindle associated protein 1 (NUSAP1)-overexpression vector indicated as NUSAP1 group, cells co-transfected with miR-758-3p mimics and NUSAP1-overexpression vector indicated as miR-758-3p mimics+ NUSAP1 group. The effects of miR-758-3p on the proliferation, migration and invasion abilities of A549 cells were detected by cell counting kit-8 (CCK-8) and Transwell assays, respectively. Bioinformatics and dual luciferase reporter assay were used to predict and verify the target gene of miR-758-3p. Results: The expressions of miR-758-3p and NUSAP1 in A549 cells were significantly up-regulated at 24 hours after transfection with miR-758-3p mimics (P<0.05). Compared with the miRNA control group (1.15±0.06), the OD value of miR-758-3p mimic group (0.78±0.06) was significantly decreased at 72 hours after transfection (P<0.05). The number of migrated cells of miR-758-3p mimic group (119.04±11.49) was significantly lower than that of the control group (271.38±19.05) (P<0.05). The number of invaded cells of miR-758-3p mimic group (71.33±5.36) was significantly lower than the control group (164.30±8.11) (P<0.05). The result of dual-luciferase reporter assay showed that NUSAP1 was a direct target of miR-758-3p. Moreover, up-regulation of NUSAP1 abolished the inhibitory effects of miR-758-3p on the proliferation, migration and invasion abilities of A549 cells. Conclusion: miR-758-3p inhibits the proliferation, migration and invasion of NSCLC cells by targeting NUSAP1.

[NUSAP1 promotes lung cancer progression by activating AKT/mTOR signaling pathway].

Objective: To investigate the inhibitory effects of nucleolar and spindle associated protein 1 (NUSAP1) on lung cancer and the related mechanisms. Methods: A549 cells were transfected with NUSAP1 siRNA, the cell proliferation, migration and invasion, and apoptosis were detected by CCK8, Transwell and flow cytometry, respectively. Western blot was used to detect the expressions of apoptosis and AKT/mTOR signal pathway related proteins. Results: Compared with the negative control group, the proliferation [(0.610±0.058) vs (1.724±0.067), P<0.05], migration [(178.267±14.780) vs (272.464±36.232), P<0.05] and invasion [(73.527±6.617) vs (120.585±13.235), P<0.05] of NUSAP1 deleted A549 cells were significantly inhibited, while the apoptosis [(3.572±0.214)% vs (11.358±1.047)%, P<0.05] was significantly increased. The expressions of apoptosis related protein Bax and active-caspase 3 were increased (P<0.05), while the expressions of anti-apoptosis protein Bcl-2 and proliferation related protein P70, the phosphorylation levels of AKT and mTOR were reduced in NUSAP1 knockdown cells (P<0.05). Conclusion: NUSAP1 knockdown can inhibit the proliferation, migration and invasion, and promote the apoptosis of tumor cells through suppressing AKT/mTOR signaling pathway in lung cancer cells.

[Expression of NUSAP1 and its relationship with prognosis in non-small cell lung cancer].

Objective: To investigate the expression of nucleolar and spindle-associated protein 1 (NUSAP1) in non-small cell lung cancer (NSCLC) and analyze its relationship with the prognosis of NSCLC patients. Methods: Real-time fluorescent quantitative PCR and immunohistochemical staining were performed to determine the expression of NUSAP1 in NSCLC tissues and adjacent tissues collected from hospital. The relationship between NUSAP1 expression and prognosis of NSCLC patients was analyzed by online database. Results: The expression level of NUSAP1 mRNA in tumor tissues was significantly higher than that of adjacent tissues (P<0.05). The high expression rate of NUSAP1 protein in NSCLC tissues was 58.0% (29/50), significantly higher than 22.0% (11/50) of adjacent tissues (P<0.05). The high expression of NUSAP1 protein in NSCLC tissues was closely correlated with tumor size, lymph node metastasis and TNM stage (P<0.05), but was not related to age and gender. The data showed that the expression level of NUSAP1 mRNA was inversely associated with the overall survival (OS) of NSCLC patients (P<0.001). The expression of NUSAP1 mRNA was significantly correlated with the pathological grade, clinical stage, gender, chemotherapy, smoking history, and histological type of NSCLC patients (P<0.05). Conclusions: The expression of NUSAP1 is up-regulated in NSCLC, which is correlated with the growth and development of NSCLC and prognosis of the patients. These results indicate that NUSAP1 can be used as a potential prognostic marker for NSCLC.

[Effect of transient ischemic attack on early neurological deterioration in patients with ipsilateral ischemic stroke].

Objective: To explore the effect of anterior circulation transient ischemic attack (TIA) on early neurological deterioration (END) in patients with ipsilateral ischemic stroke and its mechanism. Methods: (1) One hundred and thirteen patients with ipsilateral ischemic stroke and 36 healthy volunteers (healthy control group) in Neurology Department of Tianjin Medical University General Hospital from November 2014 to July 2015 were recruited into this study. According to whether got TIA before ischemic stroke, patients were divided into simple ischemic stroke group (CI group, n=87) and TIA-CI group (n=26). Their END, NIHSS score, 3-month mRS score, infarct size, serum hs-CRP and other risk factors were compared. (2) The peripheral blood mononuclear cells (PBMCs) were extracted from peripheral blood of TIA-CI group and CI group at 24 h, the 3 th day, the 7 th day and 12th day after ischemic stroke onset. At the same time, PBMCs of control group were collected. Western blot was carried out to evaluate the expression of nuclear factor-κB (NF-κB). Immunofluorescence was used to detect the cytoplasmic-to-nuclear shuttling of NF-κB. Results: (1) The incidence of END, NIHSS score at discharge, 3-month mRS score and serum hs-CRP level were significantly lower whereas the infarct size was significantly smaller of TIA-CI group than CI group (11.5% vs 31.0%, 1.9±2.3 vs 3.3±3.7, 0.9±0.8 vs 1.8±1.8, 1.1(0.3, 2.5) cm(3) vs 2.4(0.5, 22.8) cm(3,) (2.5±3.2) mg/L vs (6.2±3.2) mg/L, all P<0.05) . hs-CRP was positively correlated with END (r=0.311, P<0.05). (2) Expression of NF-κB: ① Compared with control group, the NF-κB expression increased first and then decreased in both of the two patient groups, and it decreased earlier in TIA-CI group than CI group.②In each time point, NF-κB expression of TIA-CI group was lower than CI group(t=1.754, P<0.05; t=1.858, P<0.05; t=0.609, P<0.05; t=0.519, P<0.05). (3) Activity of NF-κB: Most of NF-κB were inactivation and located in cytoplasm of control group. NF-κB of TIA-CI group and CI group was activated and translocated from cytoplasm into nuclear at the 24 h and 3 th day after ischemic stroke. At the 7 th day and 12th day, the accumulation of NF-κB in nuclear decreased and most of them located in cytoplasm in TIA-CI group, whereas most of NF-κB still located in nuclear in CI group. The activated station of NF-κB lasted shorter in TIA-CI group than CI group. Conclusions: TIA can reduce the incidence of END in patients of ipsilateral ischemic stroke. Its protective effect may relate with the inhibition of inflammation which induced by NF-κB.

[Clinical features of neurosyphilis with optic neuritis as an initial finding].

Objective: To study the characteristics of neurosyphilis with optic neuritis as an initial finding. Methods: Retrospective analysis of clinical data and laboratory testing results of 16 cases (27 eyes) with optic neuritis as an initial finding of neurosyphilis from October 2010 to March 2015 in General Hospital of People's Liberation Army was made. Results: Six-teen patients (12 males, 4 females) were collected, the median age of patients was 47 (range 33 to 65) years ,the mean age was (49.63±9.05) years. Treponema pallidum particle agglutination assay (TPPA) analysis was positive in all of the patients and rapid plasma reagin (RPR) test was positive in 14 patients (2 patients did not test). Lumbar puncture was requested and performed for all patients. Cerebrospinal fluid (CSF) TPPA analysis was positive in 16 patients and RPR test was positive in 12 patients. The CSF white blood cell counting increased in 9 (56.3%) patients and 10(62.5%)patients presented with increased CSF protein level. Both eyes were involved in 11 patients (68.8%). Relative afferent papillary defect was positive in 11 patients. Twenty-seven eyes were affected in 16 patients, and among them 7 eyes' pupil diameter were 2.5 mm or less. Incipient visual acuity was less than 0.1 in 22 eyes. The slit lamp examination showed vitreous opacity in 12 eyes and visible cells in 6 eyes among 27 eyes. Fundus examination found that 6 eyes had papillary edema and 15 eyes had pallordisc among 27eyes. Electro-retinogram (ERG) was tested in 24 eyes, and 18 eyes were abnormal. Visual evoked potential (VEP) were performed in 26 eyes (flash VEP in 22 eyes, pattern VEP in 4 eyes), and all were abnormal. Fourteen eyes were tested by 30-2 perimetry, and 6 eyes had diffuse visual field defect, 2 eyes had peripheral visual field defect, 4 eyes had quadrant defect and 2 eyes had center scotoma. Fundus fluorescence angiography was done in 16 eyes and choroidal hyper-fluorescent dots were found in posterior pole in 4 eyes. All patients were treated with antibiotic medicines, among them 10 cases in the General Hospital, and 6 cases in the other hospitals. During 15 months follow-up after discharge, visual acuity of 17 eyes recovered to 0.5 and above. Conclusion: Syphilitic optic neuritis is a condition that manifests with severe visual loss and tends to involve both eyes, Some patients have a smaller pupil diameter. Due to the particular infective routes of the disease, patients often conceal their sexual history. The manifestations of ocular syphilis are complicated and easy to misdiagnose or undiagnose. Clinical manifestations combining with the detailed history taking, serum and cerebrospinal fluid examination can guide to an accurate diagnosis and prevent from permanent vision loss. (Chin J Ophthalmol, 2016, 52: 898-904).

[Biological evaluation of three-dimensional printed co-poly lactic acid/glycolic acid/tri-calcium phosphate scaffold for bone reconstruction].

Objective: To biologically evaluate the three-dimensional(3D) printed co-poly lactic acid/glycolic acid/tri-calcium phosphate(PLGA/TCP) scaffold which could be used for repairing oral and maxillofacial bone defects, and to provide experimental evidence for its further research and clinical application. Methods: PLGA/TCP scaffolds were fabricated using low temperature rapid prototyping technique. Micro-CT and scanning electron microscope(SEM) were used to characterize the surface morphology. MC3T3-E1 cells were seeded onto the scaffold and stained with the rhodamine phalloidin and calcein acetomethoxy. After that, confocal laser scanning microscope was exploited to observe the features and viability of the cells. Moreover, the cells were co-cultured with the extract of PLGA/TCP and complete medium, respectively. The proliferation capability of the cells was assessed by the cell counting kit-8 (CCK-8) on the 1st, 2nd, and 3rd day. The PLGA/TCP scaffolds incorporated with recombinant human bone morphogenetic protein-2(rhBMP-2) of 0, 30, 60 µg(i.e. blank control group, low-dose group and high-dose group) were implanted into the latissimus dorsi muscle of the rats, and 6 weeks later, the samples were harvested to estimate the volume and pattern of new bone. Results: The 3D printed PLGA/TCP scaffold possessed a regular and well-defined porous stereo-structure with porosity of (73±3)%. Micro-CT and SEM showed that pore size were (379±32) and (453±29) µm respectively, and distance between layers were (452± 24) and (415±25) µm, and cylinder diameter were (342±24) and (350±28) µm. It also exhibited excellent cell adhesion and growth ability on the exterior and inner surface through rhodamine phalloidin and calcein acetomethoxy staining. The CCK-8 test demonstrated that the absorbance value of extract group on the 1st and 2nd day(0.51±0.08 and 0.63±0.09) were significantly higher than those in the blank control group(0.39± 0.05 and 0.53±0.05)(P<0.05), while there was no significant difference between the extract group(0.67±0.06) and the blank control group(0.68±0.04)(P>0.05) on the 3rd day. For in vivo test, there was obvious ectopic new bone formation on the PLGA/TCP scaffold incorporated with rhBMP-2, and this was demonstrated using the histological examination and micro-CT. The bone formation in the low-dose group was similar to the shape of the pre-implanted 3D printed scaffold, while much diversity was revealed in the high-dose group duo to over osteogenesis which was validated by the examinations of gross observation, histology and micro-CT. Conclusions: Customized PLGA/TCP scaffolds can be manufactured by 3D printing technique. The scaffold showed an excellent biocompatibility and ectopic osteogenesis when incorporated with rhBMP-2. However, further research is needed to validate it's effect on repairment of the oral and maxillofacial bone defects.

Mercury and cadmium contamination of irrigation water, sediment, soil and shallow groundwater in a wastewater-irrigated field in Tianjin, China.

We investigated the concentrations of Hg, Cd, Pb and As in samples of irrigation water, sediment, soil and groundwater from a field in Tianjin that was irrigated with wastewater. The results showed that the concentrations (Hg, 0.82 microg/L; Cd, 0.18 microg/L; Pb, 1.5 microg/L; As, 8.02 microg/L) in the irrigation water did not exceed the China Surface Water Quality Standard or the maximum concentrations in irrigation water recommended by the FAO. The concentrations of metals in the groundwater of wells (Hg, 0.016 microg/L; Cd, 0.128 microg/L; Pb, 0.25 microg/L; As, 4.65 microg/L) were lower than China Groundwater Quality Standard and the WHO guideline values for drinking water. The groundwater had not yet been contaminated through vertical infiltration-induced leaching. However, a substantial buildup of Hg and Cd in river sediments (I(geo) for Hg and Cd; 5.24 and 3.04, respectively) and wastewater-irrigated soils (I(geo) for Hg and Cd; 2.50 and 3.09, respectively) was observed. Taken together, these results indicated that irrigation with wastewater damaged the soil quality over the long term and that metals more easily accumulated in vegetable fields than rice fields.

[Cloning of neutral phytase gene nphy from Bacillus subtilis and its expression in Escherichia coli].

The gene encoding the neutral phytase nphy was cloned from Bacillus subtilis by polymerase chain reaction (PCR). Nucleotide sequence analysis of nphy revealed the presence of an open reading frame of 1152 bp coding for 383 aa. The start codon was followed by a sequence coding for a putative signal peptide of 26 aa in length. The nphy without original signal peptide encoding sequence was cloned into E. coli expression plasmid pTYB40. The result of SDS-PAGE of the phytase expressed in E. coli showed that the nphy had been overexpressed. The expressed phytase was over 40% of the total soluble protein of E. coli, and has normal bioactivity.

[Potentiated antitumor effect of methotrexate by dipyridamole].

Previous studies have shown that the cytotoxicity of antimetabolites to mammalian cells can be reversed by exogenous nucleosides. Dipyridamole (DP), a nucleoside transport inhibitor, can block the reversal effect, thus potentiating the cytotoxicity of antimetabolites to tumor cells. However, potentiation of antimetabolites by DP in vivo has not yet been reported. In this study we found that thymidine and hypoxanthine markedly reversed the cytotoxicity of methotrexate (MTX) to murine leukemia L1210 cells, and DP effectively blocked the reversal in vitro. In combination with amphotericin B (AmB), DP enhanced the inhibitory effect of MTX on sarcoma 180 in mice without a significant increase in toxicity. To our knowledge this is the first report that the combination of DP and AmB potentiates the antitumor effect of an antimetabolite in vivo. Results suggest that this combination may be potentially useful in cancer chemotherapy.

Potentiation of antimetabolite antitumor activity in vivo by dipyridamole and amphotericin B.

Previous studies have shown that dipyridamole (DP), a potent nucleoside transport inhibitor blocking the rescue effect of exogenous nucleosides, markedly potentiates the cytotoxicity of antimetabolites. However, no enhancement of the chemotherapeutic effect of antimetabolites by DP in vivo has yet been reported. This study provided evidence that the combination of DP and amphotericin B (AmB) significantly potentiated the inhibitory effect of 5-fluorouracil (FU) or methotrexate (MTX) against a panel of transplantable tumors including sarcoma 180, cervical carcinoma U14, and Lewis lung carcinoma in mice. No significant increase in toxicity was induced by this combination in treated mice. Our results indicate that the combination of DP and AmB with antimetabolites is potentially useful in cancer chemotherapy.