Objective:To establish the hepatic organoid of hepatitis B virus (HBV) infection on the basis of induced pluripotent stem cells (iPSC) and an inverted colloidal crystal polyethylene glycol scaffold (ICC), and to evaluate the antiviral effect of nucleoside drugs.Methods:iPSC was differentiated into hepatocyte-like cells (HLC), and inoculated into ICC to construct a hepatic organoid. The relative mRNA expressions of Nanog homeobox (NANOG), sex determining region Y-box (SOX) 2, SOX17, forkhead box protein A2 (FOXA2), alpha fetoprotein (AFP) and albumin (ALB) were detected by real time quantitative polymerase chain reaction (RT-qPCR). Confocal laser microscopy was used to photograph the three-dimension (3D) structure of organs. The expression of sodium taurocholate cotransporting polypeptide (NTCP) in HLC was analyzed by Western blot and immunofluorescence. HepG2.2.15 cells were used to extract HBV virus particles to infect hepatic organoid. The relative expression of HBV pregenome RNA (pgRNA) in cells was detected by RT-qPCR. The expressions of hepatitis B core antigen (HBcAg) and hepatitis B surface antigen (HBsAg) in cytoplasm were observed under confocal laser microscopy. A total of 0.5 μmol/L entecavir and 0.5 μmol/L lamivudine were used to treat the infected cells respectively. The relative expression of HBV pgRNA in infected and uninfected cells was detected by RT-qPCR. Independent sample t test and one-way analysis of variance were used for statistical analysis. Results:Within 21 days of iPSC differentiation, the mRNA expressions of NANOG and SOX2 in stem cells markers decreased ( F=158.90 and 8.31, respectivley; P<0.001 and P=0.002, respectively), while the mRNA expressions of SOX17 and FOXA2 in the endoderm increased first and then decreased ( F=37.23 and 82.57, respectively, both P<0.001). In the later stage of differentiation, the mRNA expressions of AFP and ALB in liver cells increased ( F=4.65 and 34.64, respectively, P=0.012 and P<0.001, respectively), and all differences were statistically significant. NTCP was highly expressed in differentiated cells detected by Western blot and fluorescence microscopy, the protein expression level was 0.803±0.099. Confocal laser microscopy confirmed that the differentiated cells expressed ALB and presented spherical structure in ICC. The expression of HBV pgRNA and the immunostaining of HBsAg and HBcAg confirmed that HBV successfully infected hepatic organoid. Three days after the application of entecavir and lamivudine, the HBV pgRNA level decreased significantly both in entecavir group (0.665±0.220) and lamivudine group (0.503±0.117) compared to the uninfected cells (3.347±0.454), and the differences were both statistically significant ( t=10.53 and 12.72, respectively, both P<0.001). Conclusions:HLC display hepatic specific genes ALB and NTCP. Hepatic organoids constructed with iPSC and ICC have human liver function and can be infected by HBV. Entecavir and lamivudine could effectively inhibit the replication of HBV in hepatic organoids.

Objective:To explore the effect and mechanism of dendritic cell derived exosomes (Dexs) loading ubiquitinated (Ub) hepatitis D antigen (HDAg) on activating specific cytotoxic T lymphocytes (CTL).Methods:Ub-S-HDAg-Dexs were co-cultured with dendritic cells (DC) which were from the femora of C57BL/6 mice for 48 h, then flow cytometry was used to detect the maturity of DC (CD86, CD80 and major histocompatibility complex (MHC) Ⅱ). The spleen-derived T lymphocytes from C57BL/6 mice were added in vitro to activate DC and co-cultivated for 72 h. The T cells were divided into Ub-S-HDAg-Dexs group (add 50 μg/mL Ub-S-HDAg-Dexs), Blank-Dexs group (add 50 μg/mL DC derived exosomes without plasmid transfection), Con-Dexs group (add 50 μg/mL DC derived exosomes transfected by cantrol plasmid), PBS group (add 50 μL/mL phosphate buffered saline), and Ub-S-HDAg-Dexs+ AG490 group (add 50 μg/mL Ub-S-HDAg-Dexs, DC and T lymphocytes stimulated by exosomes, and 50 μmol/L AG490 was also added to the cell mix). Flow cytometry was used to detect CD8 + T cells secreting interferon-gamma, non-radioactive lactate dehydrogenase release test to detect the killing activity of specific CTL. Real-time quantitative polymerase chain reaction (PCR) and Western blotting were used to detect the mRNA and protein expressions of JAK kinase (JAK) 2, GATA-binding protein 3 (GATA3), T-bet, signal transduction and activator of transcription (STAT) 1 and STAT4. Independent sample t test were used for statistical analysis. Results:The positive rates of the surface molecules CD80, CD86, MHCⅡof DC stimulated by Ub-S-HDAg-Dexs were 83.850%±0.219%、68.910%±0.134%、84.320%±0.445%, respectively.In the Ub-S-HDAg-Dexs group, the rate of CD8 + T cells secreting interferon-gamma was 6.420%±0.028%, which was higher than those of other groups, including PBS group, Blank-Dexs group, Con-Dexs group and Ub-S-HDAg-Dexs+ AG490 group ( t=90.78, 30.32, 63.06 and 85.42, respectively, all P<0.001). The cytotoxicity of T cells in the Ub-S-HDAg-Dexs group was 82.4%±3.9%, which was higher than those of other groups ( t=17.28, 9.74, 3.95 and 15.89, respectively, all P<0.050). The relative mRNA expressions of JAK2, T-bet, STAT1, STAT4 in Ub-S-HDAg-Dexs group were higher than those in other groups, including Con-Dexs group ( t=10.74, 32.34, 13.00 and 16.28, respectively, all P<0.001), Blank-Dexs group ( t=15.05, 21.51, 6.46 and 13.12, respectively, all P<0.050), PBS group ( t=21.83, 41.42, 7.30 and 17.50, respectively, all P<0.050), Ub-S-HDAg-Dexs+ AG490 group ( t=35.75, 20.69, 17.02 and 17.07, respectively, all P<0.001), and the differences were all statistically significant. The protein expressions of T-bet, STAT1, STAT4 in Ub-S-HDAg-Dexs group increased compared with those in PBS group ( t=346.70, 57.54 and 55.81, respectively, all P<0.001), with statistical significance. In the presence of AG490, the protein expressions of T-bet, STAT1 and STAT4 decreased compared with those in Ub-S-HDAg-Dexs group, and the differences were statistically significant ( t=355.40, 52.79 and 126.10, respectively, all P<0.001). Conclusions:Ubiquitinated HDAg transported by exosomes could effectively promote DC maturation, induce T lymphocyte differentiation, and generate specific CTL responses, which provides a new idea for the treatment of hepatitis D.

Objective:To investigate the effects of antibiotic treatment and antibiotics combined with surgery treatment on the prognosis of patients with infective endocarditis (IE).Methods:The clinical data and prognosis of all patients diagnosed as IE discharged from Shanghai Jiao Tong University Affiliated Sixth People′s Hospital from June 2011 to May 2021 were collected. There were 240 IE patients, divided into antibiotic treatment group and the antibiotics combined with surgery group according to the treatment methods. The clinical characteristics and prognosis of the IE patients were compared between the two groups, so as to investigate the timing of surgery for IE patients and to analyze the effects of the two treatment methods on the prognosis of IE patients.Statistical analysis methods including Wilcoxon rank sum test, chi-square test, Kaplan-Meier survival analysis and Cox regression analysis were used when appropriate.Results:Of the 240 patients with IE, 63 cases were only treated with antibiotics and 177 cases were treated with antibiotics combined with surgery. After propensity score matching (PSM), one-year mortality rate of the IE patients in the antibiotics combined with surgery group was 11.1%(4/36), which was significantly lower than that in the antibiotic treatment group (33.3%(12/36), χ2=5.14, P=0.023). The median values of left ventricular ejection fraction (LVEF), left ventricular end diastolic diameter (LVEDD) and left ventricular fractional shortening (LVFS) in the antibiotics combined with surgery group were 59%, 47 mm and 31%, respectively, which were significantly lower than those before surgery (63%, 54 mm and 34%, respectively, Z=6.19, 9.36 and 6.11, respectively, all P<0.001). The most common surgical indication was moderate to severe heart failure, and there was no significant difference between the early operation group and the late operation group (both P>0.050). The one-year cumulative survival rate of antibiotics combined with surgery group was 94.9%, which was significantly higher than that in the antibiotic treatment group (83.2%, χ2=7.38, P=0.007). Heart failure and Pitt bacteremia scores≥4 were the independent risk factors for one-year all-cause death of the IE patients (hazard ratio ( HR)=5.668 and 19.392, respectively, both P<0.050). Hospital days and antibiotics combined with surgery were independent related factors for reducing the risks of one-year all-cause death ( HR=0.931 and 0.299, respectively, both P<0.050). Pitt bacteremia scores≥4 had the greatest impact on one-year prognosis of the IE patients. Conclusions:Surgery could significantly improve cardiac function and one-year prognosis of the IE patients. IE patients with heart failure and Pitt bacteremia score≥4 should be actively treated.

Lymphocytic hypophysitis(LYH) is a rare autoimmune disease, which is more common in women and is closely associated with pregnancy. For patients with lymphocytic hypophysitis, the clinical manifestations mainly depend on the speed of disease progression, the extent of lesions, the severity and stage of disease, which can present as mass effect, decreased anterior pituitary hormone function, and central diabetes insipidus, etc. Due to physical changes in pituitary anatomy and hormone levels, as well as restriction of imaging examinations during pregnancy, it poses greater challenge to establish diagnosis in perinatal patients, which demands a comprehensive assessment of the patient′s clinical symptoms and biochemical tests. For treatment, patients with hypopituitarism during pregnancy should be given hormone replacement therapy. Moreover, patients need to adopt optimized tailored therapy according to gestational age and various clinical manifestations. At present, there is no report on the adverse effects of LYH on pregnancy outcome and re-pregnancy. This review discusses the diagnosis, treatment and prognosis of patients with LYH during pregnancy, and provides reference for the clinical management of this condition.

Objective:To clarify the effect of ubiquitination hepatitis B virus core antigen (Ub-HBcAg) on dendritic cells (DC) autophagy, and to explore the mechanism of autophagy in enhancing DC antigen presentation and inducing hepatitis B virus-specific cytotoxic T lymphocyte (CTL) responses.Methods:Ub-HBcAg lentiviral vector (LV-Ub-HBcAg), lentiviral vector-hepatitis B virus core antigen (LV-HBcAg) and no-load plasmid LV (LV) were constructed and packaged. DC2.4 cells were divided into LV-Ub-HBcAg group, LV-HBcAg group and LV group. The blank control group (NC group) was also set. The protein expression of autophagy-related protein P62, microtubule associated protein 1 light chain 3 beta (LC3B), autophagy related 5(ATG5) and Beclin-1 were detected by Western blotting. The expressions of co-stimulatory molecules such as CD86, CD80 and major histocompatibility complex (MHC)-Ⅱ were detected by flow cytometry. Cell counting kit-8 (CCK-8) method was used to detect T lymphocytes proliferation. The non-radioactive lactic acid dehydrogenase (LDH) release method was applied to detect the killing ability of CTL. Statistical analysis was conducted by independent sample t test. Results:The relative protein expressions of LC3B-Ⅱ/LC3B-Ⅰ, Beclin-1 and ATG5 in NC group were 0.445±0.076, 0.522±0.026 and 0.761±0.038, respectively, which were all lower than those in LV-Ub-HBcAg group (0.926±0.021, 0.919±0.016 and 1.451±0.028, respectively). The relative protein expression of P62 in the NC group was higher than that in LV-Ub-HBcAg group ((1.875±0.016) vs (0.647±0.121)). The differences were all statistically significant ( t=6.102, 9.842, 17.490 and 10.590, respectively, all P<0.01). The expressions of CD86 (75.51%), CD80 (83.35%), MHC-Ⅱ (66.66%) in the LV-Ub-HBcAg group were high, and those in the NC group were 8.03%, 7.49%, 0.04%, respectively. The specific CTL killing rate ((65.310±2.091)%) of the LV-Ub-HBcAg group was significantly higher than both NC group ((14.400±0.497)%) and LV-HBcAg group ((54.870±1.443)%), and the differences were both statistically significant ( t=23.690 and 4.111, respectively, both P<0.05). Conclusion:Ub-HBcAg promotes the DC autophagy, up-regulates the expressions of costimulatory molecules on cell surface of DC to induce the maturation and activation, and then stimulates T lymphocyte to induce a stronger specific CTL response under the effort of ubiquitination.

Objective:To analyze the clinical characteristics of fulminant type 1 diabetes (FT1DM) in China.Methods:Clinical data of 279 cases related to FT1DM in Chinese Database from January 2005 to December 2018 were collected, and other 20 patients from our hospital were included in the present study.Results:(1) There has been a progressive increasing in the number of reported cases every year in China, and the number in the southern region were significantly more than that in the northern region. (2) The median age of the onset of FT1DM patients in China was 32.5 years old, without significant gender difference. Moreover, 36.5% (54/148) of the female patients caught the disease during their prenatal period, most of them were onset in the second or third trimesters of pregnancy and 2 weeks after delivery (37/40), and the prognosis of the fetus was extremely poor. (3) Compared with new-onset type 1 diabetes, FT1DM patients were younger, and with higher blood glucose [(39.7±15.3) vs (21.2 ± 9.9) mmol/L], higher serum creatinine [(188.4±115.9) vs (51.8 ±23.1) μmol/L], and higher amylase levels [245.5 (26.0-5 137.0) vs 54.7 (14.0-404.9) U/L]. FT1DM patients were with more severe acidosis, and lower HbA 1C level [(6.6 ±0.8)% vs (12.9 ± 2.5)%, P<0.01]. (4) FT1DM patients may combine with multiple organ dysfunction or severe metabolic disorders, electrolyte disorders, as well as liver and kidney dysfunctions, and elevation of amylase and muscle enzymes. Conclusion:FT1DM are with some clinical characteristics different from classic new-onset type 1 diabetes, including adult-onset, frequent in the southern China. Pregnancy may be a predisposing factor for female patients. Significant metabolic disorders and multiple organ involvements are common in the patients with FT1DM.

This article reported a case of malignant pheochromocytoma and paraganglioma with cranial metastasis. The patient was a 25-year-old man. At the age of 10, he was diagnosed with retroperitoneal paraganglioma. He received radiotherapy after partial resection of the tumor. At the age of 15, when he consulted for headache and hypertension, a space occupying lesion was found on the right adrenal without further diagnosis and treatment. His headache and hypertension were aggravated at the age of 25. Then he visited our medical center and diagnosed with right adrenal pheochromocytoma. He undertook an operation to remove the right adrenal lesion, and his blood pressured could be controlled in the normal range with oral antihypertensive medication. One year after the surgery, a cranial lesion with positive 131I-metaiodobenzylguanidine imaging was found following the increase of norepinephrine and dopamine in both blood and urine. The diagnosis of malignant pheochromocytoma with cranial metastasis was confirmed. The cranial lesion was treated with cyberknife therapy for 4 times, and the lesion was slightly reduced. Four year after the removal of right adrenal tumor, he developed multiple bone metastasis in the right ribs and left ilium, and then he received 131I-metaiodobenzylguanidine radiotherapy. Additionally, the results of gene test indicated a pathogenic mutation of SDHB. During the follow-up period of one year after the treatment, his blood pressure fluctuated to about 140/90mmHg with 10mg of phenoxybenzamine once a day. The case is a malignant pheochromocytoma with multiple bone metastasis characterized by a long course with an ideal prognosis in a way. Consequently, we suggested the diagnostic pathway with genetic testing based on clinical phenotype to approach more timely diagnosis and reasonable evaluation of the prognosis in pheochromocytoma and paraganglioma. Furthermore, clinicians should pay more attention to the long-term surveillance.

Objective To determine the correlation between papillary thyroid cancer( PTC) malignancy with thyroid antibody expression level. Methods In this retrospective study, 4046 goiter patients with complete clinical data who have undergone thyroidectomy were separated into the benign nodule ( n=1357) and PTC groups ( n=2689), and their preoperative thyroglobulin antibody ( TgAb) and thyroid peroxidase antibody ( TPOAb) were assessed through antibody analysis. HT was determined based on the postoperative histopathology. The collected data were then evaluated for the correlation with PTC malignancy through statistical tests. Results In general, patients in the PTC group showed a significantly higher TgAb positivity ( 10. 9％) than those in the benign group ( 5. 6％, P<0.01) . Among female patients, a higher TgAb positivity was observed for those in the PTC group than those in the benign group (14.0％ vs 7.0％, P<0.01), while no significant difference between groups was observed in male patients. The high odds ratio of 2.18 supports the significant correlation between TgAb-positive status and higher risk of developing PTC in women (P<0.05). However, the risk of developing PTC was not correlated with higher TgAb titre. TgAb was not significantly correlated with the mass size, multifocality, lymph node metastasis or the TNM classification of malignant tumor stage Ⅲ/Ⅳ. Conclusion The increasing risk of PTC is related to TgAb among female patients, but it will not increase as TgAb titer rises. It is not associated with PTC aggressiveness.

Cyclospora cayetanensis is a foodborne and waterborne pathogen that causes endemic and epidemic human diarrhea worldwide. A few epidemiological studies regarding C. cayetanensis infections in China have been conducted. During 2013, a total of 291 stool specimens were collected from patients with diarrhea at a hospital in urban Shanghai. C. cayetanensis was not detected in any of the stool specimens by traditional microscopy, whereas five stool specimens (1.72%, 5/291) were positive by PCR. These positive cases confirmed by molecular technology were all in the adult group (mean age 27.8 years; 2.94%, 5/170) with watery diarrhea. Marked infection occurred in the rainy season of May and July. Sequence and phylogenetic analyses of the partial 18S rRNA genes of C. cayetanensis isolated showed intra-species diversity of this parasite. This study showed, for the first time, that C. cayetanensis is a pathogen in outpatients with diarrhea in Shanghai, albeit at a low level. However, the transmission dynamics of this parasite in these patients remain uncertain.
Adolescent ; Adult ; China ; epidemiology ; Cyclospora ; genetics ; isolation & purification ; Cyclosporiasis ; epidemiology ; Diarrhea ; etiology ; parasitology ; Feces ; parasitology ; Female ; Humans ; Male ; Middle Aged ; Outpatients ; Phylogeny ; Polymerase Chain Reaction ; RNA, Ribosomal, 18S ; analysis ; Retrospective Studies ; Young Adult

Objective To investigate the characteristics of peroperative blood pressure and heart rate in patients with normotensive incidental pheochromocytomas in order to provide the basis for peroperative treatment. Methods This retrospective study collected the data of 104 patients with a pathological diagnosis of unilateral pheochromocytoma at PLA General Hospital during January 2011 to December 2016. They were divided into normotensive incidental pheochromocytomas(NIP) group (n＝50) if the patients were normotensive and HIP group ( n＝54) if the patients were with hypertension. The clinical features, imaging features and peroperative hemodynamics were analyzed. Results ( 1) The age, urinary norepinephrine, daily dosage and duration of phenoxybenzamine in NIP group were less than those of HIP group (all P<0.05). (2) Preinduction blood pressure, maximum blood pressure, and total fluid intake in NIP group were lower than those in HIP group(all P<0.05). The blood pressure range, heart rate range, increased blood pressure, minimum mean arterial pressure, vasoactive medication were without statistical significance between these two groups. ( 3) The times and rate of intraoperative systolic blood pressure more than 30% baseline, 200 mmHg (1 mmHg＝0.133 kPa), 180 mmHg, 160 mmHg, intraoperative tachycardia, bradycardia, intraoperative hypotension and postoperative hypotension were without statistical significance between these two groups. (4) Stratified analysis of age (50 years), phenoxybenzamine (40 mg/d), tumor diameter (50 mm) and preinduction blood pressure (130/80 mmHg) showed that intraoperative blood pressure and heart rate were without statistical significance between these two groups. ( 5) There was no correlation between phenoxybenzamine ( daily dosage or duration ) and peroperative hypotension. Applying phenoxybenzamine or vasoactive medication was not correlated with peroperative hypotension in NIP group. Conclusion The peroperative blood pressure and heart rate of patients with NIP are similar to those of patients with HIP. Adequate peroperative treatment should be applied to NIP to avoid hemodynamic instability.