[Study on the resistance of rifampicin-resistant Mycobacterium tuberculosis to anti-tuberculosis drugs in group A].

Objective: To summarize the resistance of rifampicin-resistant Mycobacterium tuberculosis to anti-tuberculosis drugs in group A. Methods: In the retrospective study, a total of 1 226 clinical isolates from suspected multidrug-resistant pulmonary tuberculosis patients in Beijing TB control system from 2016 to 2021 were identified as Mycobacterium tuberculosis (MTB) strains by MPB64 antigen detection test. Rifampicin-resistant tuberculosis (RR-TB) strains were screened by the phenotypic drug susceptibility using the proportion method. The drug susceptibilities of Levofloxacin(LFX), Moxifloxacin(MFX), Bedaquiline(BDQ) and Linezolid(LZD)were detected by the phenotypic drug susceptibility with microplate method. The drug resistance rate, drug resistance level and minimum inhibitory concentration (MIC) distribution of four anti-tuberculosis drugs in group A were analyzed. We calculated the demographic distribution of RR-TB, multidrug-resistant tuberculosis(MDR-TB), pre-extensively drug resistant tuberculosis (pre-XDR-TB), extensively drug resistant tuberculosis (XDR-TB) patients and the cross resistance of LFX and MFX, then summarized the drug-resistance spectrum of BDQ-resistant and LZD-resistant strains and the treatment outcome of RR-TB patients. Measurement data were expressed as rate or composition ratio,χ2 test was used between and within groups, and P<0.05 was considered statistically significant. Results: Among the 1 226 suspected multidrug-resistant pulmonary tuberculosis patients, the detection rates of RR/MDR/pre-XDR/XDR-TB patients were 20.8%(255/1 226), 15.2%(186/1 226), 5.7%(70/1 226), 0.5%(6/1 226), respectively. There were statistically significant differences in the distribution of patients with the four types of drug resistance in terms of age and treatment history (χ2=14.95, P=0.020;χ2=15.91, P=0.001). The drug resistance rates of LFX, MFX, BDQ and LZD in RR-TB patients were 27.5% (70/255), 27.5% (70/255), 0.4% (1/255) and 2.4% (6/255), respectively. The MICs of LFX, MFX and LZD-susceptible MTB were mainly at 0.25 mg/L, and the MIC of BDQ-susceptible MTB was mainly concentrated at 0.03 mg/L. 25.1% (64/255) of the RR MTB were resistant to both LFX and MFX, and 6 strains were resistant to LFX or MFX, showing incomplete two-way cross resistance. One BDQ-resistant strain and six LZD-resistant strains were detected. The treatment success rate of RR-TB patients was 74.4% (151/203), and there were statistically significant differences in treatment outcomes between resistant and sensitive patients on the LFX-containing treatment regimen (Fisher's exact test, P=0.012). Conclusions: The prevalence of fluoroquinolones (LFX and MFX) resistance in rifampicin-resistant MTB is very serious. LFX and MFX show incomplete bidirectional cross-resistance. BDQ and LZD have the most promising future in the treatment of MDR-TB. Improve drug-resistance testing will help to further improve the success rate of treatment.

[Ferroptosis in laryngeal squamous cell carcinoma and its regulation by M2 macrophage-derived exosomes].

Objective: To investigate ferroptosis in laryngeal squamous cell carcinoma (LSCC) and its regulation by M2 macrophage-derived exosomes. Methods: LSCC and adjacent noncancerous tissue samples were collected from 32 patients treated in the Department of Otorhinolaryngology, Head and Neck Surgery of the Second Affiliated Hospital of Harbin between September 2018 and April 2021, including 26 males and 6 females, aged 43-79 years. The expressions of ferroptosis marker glutathione peroxidase 4(GPX4) in LSCC and adjacent noncancerous tissues were detected by immunohistochemistry and reverse transcriptase-polymerase chain reaction(RT-PCR). The correlations between GPX4 expression and clinicopathological factors in LSCC were analyzed. Biological changes of TU212 cells after treated with ferroptosis-induced agent erastin were detected by transmission electron microscope, cell counting kit-8(CCK-8), clone test, reactive oxygen species(ROS), malondialdehyde(MDA), glutathione(GSH), JC-1, RT-PCR and western blot. Exosomes were isolated from the supernatant of M0/M2 macrophages (M0-exos/M2-exos) and co-incubated with erastin-treated TU212 cells to detect the change of ferroptosis in cells of each group. The data were analyzed by SPSS software of version19.0. Results: GPX4 expression in LSCC tissues was significantly higher than that in adjacent noncancerous tissues (2.04±0.65 vs. 0.99±0.09, F=30.36, P<0.001), and was closely related to T stage and clinical stage (Ⅰ-Ⅱvs.Ⅲ-Ⅳ: 1.75±0.39 vs. 2.18±0.71, F=2.25, P<0.05; T1-2 vs. T3-4: 1.71±0.42 vs. 2.20±0.69, F=2.06, P<0.05). In TU212 cells treated with erastin, mitochondrial crest became smaller, membrane density increased, proliferation rate decreased, intracellular ROS level increased, mitochondrial membrane potential depolarized, GSH content decreased, intracellular MDA level increased and expressions of GPX4 mRNA and protein decreased. Change of M0 into M2 macrophages was induced by IL-4 stimulation. When erastin-treated TU212 cells were incubated with M2-exos, cell proliferation was partially restored and GPX4 expression was enhanced, and also with the recoveries of levels of ROS, MDA and GSH (all P<0.05). Conclusions: Ferroptosis is one of the cell death ways of LSCC. M2-exos may inhibit ferroptosis of LSCC cells.

[The function and mechanism of long non-coding RNA RP11-159K7.2 in sinonasal squamous cell carcinoma].

Objective: To explore the role and mechanism of long non-coding RNA RP11-159K7.2 in the progression of sinonasal squamous cell carcinoma (SNSCC). Methods: Sixty-five cases of SNSCC tissues and adjacent tissues were selected from the Department of Otorhinolaryngology Head and Neck Surgery, the Second Affiliated Hospital of Harbin Medical University from 2009 to 2014. The expression of RP11-159K7.2 in SNSCC and adjacent tissues was detected by RNAscope in situ hybridization to observe its association with prognosis. Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR associated proteins 9 (CRISPR/Cas9) was used to knockout the expression of RP11-159K7.2 in RPMI-2650 cells (SNSCC cell line). Cell counting kit-8 (CCK-8), wound healing and Transwell were performed to observe the changes of proliferation, migration and invasion of SNSCC cells in vitro after down-regulation of RP11-159K7.2. Moreover, the growth of xenograft in nude mice after down-regulation of RP11-159K7.2 was examined in vivo. Mechanically, the protein chip, Western blot and RNA immunoprecipitation were performed to identify the proteins bound by RP11-159K7.2. SPSS 17.0 was used for statistical analysis. Results: The expression of RP11-159K7.2 in SNSCC tissue was significantly higher than that in adjacent tissues. RP11-159K7.2 expression was closely related with T grade, nodal metastasis and differentiation of SNSCC (χ2 value was 4.697, 4.235 and 10.753, respectively, all P<0.05). The five-year survival rate of RP11-159K7.2 high expression patients was significantly lower than that of RP11-159K7.2 low expression ones (P=0.013 7). After the down-regulation of RP11-159K7.2, the proliferation, migration and invasion ability of SNSCC cells decreased significantly, and the growth of SNSCC xenograft was significantly inhibited. There were 31 candidate proteins that may bind to RP11-159K7.2. RP11-159K7.2 directly bound to nuclear factor-κB (NF-κB) in SNSCC cells, and the regulation of RP11-159K7.2 on the proliferation and invasion of SNSCC cells depended on NF-κB. Conclusion: The increased expression of RP11-159K7.2 in SNSCC may serve as a potential molecular marker for SNSCC prognosis assessment. It is currently considered that the carcinogenic mechanism of RP11-159K7.2 in SNSCC is related to the regulation of NF-κB protein.

[Application of rigid curved laryngoscope in the examination of hypopharynx and the treatment of early hypopharyngeal cancer].

Objective: To evaluate the efficacy of curved suspension laryngoscope assistant transoral surgery (CLATOS) in the examination of hypopharynx, and diagnosis and surgery of early hypopharyngeal carcinoma. Methods: Thirty-four patients who underwent detailed examination for lower part of hypopharynx and surgery for early hypopharyngeal carcinoma with CLATOS technique in the Second Affiliated Hospital of Harbin Medical University from January 2019 to January 2020 were analyzed retrospectively. The age ranged from 42 to 74 years old including 28 males and 6 females. Among them, 25 patients complaining of foreign body sensation in the pharynx for more than half a year with a poor exposure of the lower pharynx in the examination with flexible laryngoscope in the outpatient department were admitted to the hospital for the further examination with rigid curved laryngoscopy and 9 patients with stage T1-2 hypopharyngeal squamous cell carcinoma were operated with CLATOS technique. Preoperative, intraoperative and postoperative data were analized. Results: One case of squamous cell carcinoma in esophageal entrance (T1N0M0) and one case of carcinoma in pyriform sinus (T1N0M0) with cervical esophageal carcinoma (T1N0M0) were found in the 25 patients with foreign body sensation in the pharynx. En bloc resection of cancer was obtained in 9 patients with stage T1-2 hypopharyngeal carcinomas and 2 of them underwent tracheotomy. One patient with T1 retrocricoid carcinoma was found to have a carcinoma in situ at the lower part of posterior hypopharyngeal wall in the examination during follow-up, which was resected simultaneously. Postoperatively this patient developed a stenosis in the esophageal entrance, which was dilated twice until swallowing normally. Two patients of T2N0M0 and one of T2N1M0 underwent bilateral neck lymph node dissection just after the removal of primary tumors. During follow-up, none of the 9 patients showed tumor recurrence and complications such as dysphonia and dyspnea. Conclusion: CLATOS technique can provide a promising way in the examination of hypopharynx and the detection of early lesions of hypopharynx and esophageal entrance, and this technique has also the advantages such as full visual angle and easy operation for the resection of early hypopharyngeal carcinoma. The quality of life of patients can be improved while the tumor is removed en bloc with this technique.

KLF15 reduces the level of apoptosis in mouse liver induced by sepsis by inhibiting p38MAPK/ERK1/2 signaling pathway.

OBJECTIVE: Sepsis-induced acute liver injury (ALI) involves multiple systems in the body. The disease is acute and critical, with various symptoms, including extensive necrosis of liver cells. There is currently no effective treatment to deal with ALI in a timely manner. This study verified the therapeutic effect of Krüppel-like factor 15 (KLF15) on ALI by studying its anti-apoptotic effect on the liver. MATERIALS AND METHODS: We induced ALI in mice with lipopolysaccharide (LPS)/D-galactosamine (D-GaIN). Recombinant mouse KLF15 was used to treat mice to examine the protective effects of KLF15 on mouse liver and the effects of apoptosis-related molecules. In addition, we cultured Kupffer cells and determined the anti-inflammatory and anti-apoptotic effects of KLF15 and its mechanism by overexpressing KLF15. RESULTS: Exogenous KLF15 effectively reduced the levels of TIBL, ALT, AST, and inflammatory factors (COX-2, MCP-1, IL-1ß, and TNF-α) in mouse serum. The results of HE staining also demonstrate that KLF15 improves the morphology of liver tissue. In addition, the expression of KLF15 in LPS-induced Kupffer cells was significantly reduced and KLF15 increased the viability of Kupffer cells and decreased the level of inflammation in Kupffer cells. In both in vivo and in vitro experiments, KLF15 reduced the level of apoptosis in hepatocytes or Kupffer cells and inhibited the activity of the p38MAPK/ERK1/2 signaling pathway. CONCLUSIONS: KLF15 reduces the apoptosis and inflammation levels of liver and Kupffer cells by inhibiting the p38MAPK/ERK1/2 signaling pathway and alleviates LPS/D-GaIN-induced ALI.

Nesfatin-1 alleviates acute lung injury through reducing inflammation and oxidative stress via the regulation of HMGB1.

OBJECTIVE: Acute lung injury (ALI) is the most common complication of sepsis, with rapid onset and high mortality. There is currently no effective treatment for ALI. Therefore, we looked for a good method of treating ALI by studying the effect and mechanism of Nesfatin-1 on ALI. MATERIALS AND METHODS: We used LPS to induce mouse and human alveolar epithelial cell line BEAS-2B to construct an ALI model. Recombinant Nesfatin-1 was administered subcutaneously to mice or used to stimulate BEAS-2B cells. We collected mouse bronchoalveolar lavage fluid and mouse lung tissue to detect changes in inflammatory factors and oxidative stress levels. In addition, we examined the expression changes of HMGB1 to study the mechanism of Nesfatin-1. RESULTS: Exogenous Nesfatin-1 significantly attenuated LPS-induced ALI and reduced inflammation levels and oxidative stress levels in mouse lung tissue. In cell experiments, Nesfatin-1 also reduced inflammation levels and oxidative stress levels in BEAS-2B cells. In addition, Nesfatin-1 reduced the expression of HMGB1 in mouse lung tissues and BEAS-2B cells, and decreased the activity of p38MAPK and NF-κB signaling pathways in the inflammation-related pathway downstream of HMGB1. However, after overexpression of HMGB1, the therapeutic effect of Nesfatin-1 on ALI was attenuated. CONCLUSIONS: Nesfatin-1 regulates the expression of HMGB1 in alveolar epithelial cells. By reducing the expression of HMGB1, Nesfatin-1 can reduce the inflammation-related signaling pathway downstream of HMGB1 to reduce the level of inflammation and oxidative stress in alveolar epithelial cells, thereby alleviating ALI.

[Comprehensive analysis of the effect of null zone shifting surgery treatment on patients with infantile nystagmus syndrome].

Objective: To characterize the postoperative change of eyes related parameters of patients with infantile nystagmus syndrome(INS), so as to provide a reference for the clinical evaluation of postoperative effect and the rational arrangement of patients' follow-up time after operation. Methods: A retrospective study. Clinical and follow-up data of 17 patients diagnosed with INS at Department of Ophthalmology in Xinhua Hospital, School of Medicine, Shanghai Jiaotong University from June 2014 to December 2016 were collected. All patients with abnormal head posture (AHP) underwent null zone shift surgery. The operative methods were Parks 5-6-7-8, Anderson, Kestenbum 5-5-6-4,null zone shift combined with strabismus correction and vertical null zone transposition. Ophthalmological examination and eye movement were recorded, including best corrected binocular visual acuity (BCBVA), position of the null zone, expanded nystagmus acuity function (NAFX) and foveation time. Single factor repeated analysis of variance, independent sample t test and rank sum test were used for statistical analysis. Results: Among the 17 children, 6 were females and 11 were males. The age at surgery of the patients was 5-11 years.The follow-up time was (14.8±6.0) months. Preoperative BCBVA was 0.382±0.147 (corrected posture), 0.300±0.056 (AHP); foveation time was (0.594±0.011)s;position of null zone was 23.570°±0.118°. The BCBVA at three months after operation was 0.318±0.044 (corrected posture), 0.260±0.045 (AHP); foveation time was (0.950±0.146)s; position of null zone was 5.360°±1.107°. The BCBVA at six months after operation was 0.264±0.039 (corrected posture), 0.230±0.037 (AHP);foveation time was (1.496±0.233) s; position of null zone was 6.070°±1.303°. The BCBVA at twelve months after operation was 0.309±0.039 (corrected posture), 0.250±0.045 (AHP);foveation time was (1.455±0.201) s; position of null zone was 9.290°±8.520°. There was statistical difference between the data of pre-operation and post-operation(all P<0.05). Change of null zone position was identified in six patients after six months.The preoperative NAFX of patients with presence of change of null zone positon was 0.308±0.063 (the primary position), 0.393±0.210 (null zone); BCBVA was 0.450±0.043 (corrected posture), 0.417±0.031 (AHP); foveation time was 0.122 (0.080-1.014)s. The postoperative NAFX of those patients was 0.430±0.090(the primary position), 0.471±0.140 (null zone); foveation time was 0.438(0.170-1.450) s. The data above were lower than that of patients with no regression of null zone[0.523±0.142,0.601±0.110,0.200±0.063,0.250±0.076,0.725(0.230-1.440)s,0.610±0.160,0.680±0.120,0.975(0.380-2.000)s]. The difference was statistically significant(all P<0.05). Conclusions: Null zone shifting surgery is an effective approach for treating INS. Reduction in the null zone position can be observed in some patients at 6 months after operation, which was related to NAFX, BCBVA and foveation time. It is recommended to extend the follow-up time to at least 6 months after the operation. (Chin J Ophthalmol, 2019, 55:13-19).

[Establishment of Automation System for Detection of Alcohol in Blood].

OBJECTIVES: To establish an automation system for detection of alcohol content in blood. METHODS: The determination was performed by automated workstation of extraction-headspace gas chromatography （HS-GC）. The blood collection with negative pressure, sealing time of headspace bottle and sample needle were checked and optimized in the abstraction of automation system. The automatic sampling was compared with the manual sampling. RESULTS: The quantitative data obtained by the automated workstation of extraction-HS-GC for alcohol was stable. The relative differences of two parallel samples were less than 5%. The automated extraction was superior to the manual extraction. A good linear relationship was obtained at the alcohol concentration range of 0.1-3.0 mg/mL （r≥0.999） with good repeatability. CONCLUSIONS: The method is simple and quick, with more standard experiment process and accurate experimental data. It eliminates the error from the experimenter and has good repeatability, which can be applied to the qualitative and quantitative detections of alcohol in blood.

[The first confirmed imported case of yellow fever in China].

OBJECTIVE: To describe the epidemiological characteristics of the first confirmed imported case of yellow fever in China. METHODS: This case was reported through the Infectious Disease Surveillance Program. Information on epidemiology and clinical manifestation of the case was collected through case interview and related medical records. Blood and saliva samples of the case were collected and tested by real-time PCR. RESULTS: The patient was male, 32 years old, and suffered a sudden onset of fever without other symptoms, on March 8(th), 2016. The patient arrived in Beijing at midnight on March 10(th). Condition of the patient got progressively worsened, with both liver and renal failures, hepatic encephalopathy, multiple organ failures and DIC, finally died on March 16(th). Serum of the case was positive for yellow fever virus by real time PCR. The patient was bit by mosquitoes six days before the onset of fever in Luanda, Angola. CONCLUSION: This report summarized related information on the first confirmed but imported case of yellow fever in China that was helpful to the management of other imported yellow fever cases in the future.

Dopamine pathway loss in nucleus accumbens and ventral tegmental area predicts apathetic behavior in MPTP-lesioned monkeys.

Apathy, primarily defined as a lack of motivation, commonly occurs in people with Parkinson disease (PD). Although dysfunction of basal ganglia pathways may contribute to apathy, the role of dopamine remains largely unknown. We investigated the role of dopaminergic pathways in the manifestation of apathetic behaviors by measuring the effects of the selective dopaminergic neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) on monkeys' willingness to attempt goal directed behaviors, distinct from their ability to perform tasks. Fifteen macaques received variable doses of MPTP, had PET scans with [(11)C]-dihydrotetrabenazine (DTBZ), [(11)C]-2ß-3ß-4-fluorophenyltropane (CFT), and [(18)F]-fluorodopa (FD) and performed tasks to assess apathetic behaviors and motor impairment. At 8 weeks post-MPTP, primates were euthanized and stereological cell counts and dopamine measurements were done. Apathy scores were compared to motor scores, in vitro and in vivo dopaminergic measures. Apathy scores increased following MPTP and correlated with DTBZ (r(S) = -0.85), CFT (r(S) = -0.87), and FD (r(S) = -0.85) specific uptake in nucleus accumbens (NAcc,) and dopaminergic cell counts in ventral tegmental area (VTA, r(S) = -0.80). Dopaminergic cell loss in VTA provided significant predictive power for apathy scores after controlling for the influence of cell loss in SN. Additionally, forward step-wise regression analyses indicated that neuropathological changes in the VTA-NAcc pathway predict apathetic behavior better than motor impairment or neuropathological changes in the nigrostriatal network. Our findings suggest that dopaminergic dysfunction within the VTA-NAcc pathway plays a role in the manifestation of apathetic behaviors in MPTP-lesioned primates. Similar changes in people with PD may contribute to apathy.

Serological survey of 2009 H1N1 influenza in residents of Beijing, China.

In order to determine the prevalence of antibody against 2009 H1N1 influenza in Beijing, we conducted a serological survey in 710 subjects, 1 month after the epidemic peak. We found that 13·8% of our cohort was seropositive. Subjects aged ≥60 years recorded the lowest seroprevalence (4·5%). The age-weighted seroprevalence of 14·0% was far lower than the supposed infection rate at the epidemic peak, derived from the basic reproduction number for 2009 H1N1 virus. For subjects who had received the pandemic vaccine seroprevalence was 51·4%. In subjects aged ≥60 years the seasonal influenza vaccination was not significantly associated with being seropositive. Our study suggests that many factors, and not just the immunological level against 2009 H1N1 influenza in the community, affected the spread of the virus within the population of Beijing.