Structural mechanism for multidrug efflux systems

Bacteria such as Escherichia coli, Campylobacter jejuni and Neisseria gonorrhoeae have developed various mechanisms to overcome toxic environments that are otherwise unfavorable for their survival. One important strategy that bacteria use to expel toxic compounds, including heavy metal ions, is the expression of membrane efflux transporters that recognize and actively export these toxic compounds out of bacterial cells, thereby allowing them to survive in extremely toxic conditions. Many of these transporters are multiple drug binding proteins which extrude different toxic chemicals and mediate a phenomenon of multidrug resistance (MDR) in bacteria. The expression of these efflux transporters is tightly controlled at the transcriptional level by transcriptional regulators. A number of these transcriptional regulators are also multidrug binding proteins, which recognize and respond to the same set of toxic chemicals that are expelled by the efflux transporters they regulate. The goal of this dissertation is to elucidate the structures and fundamental mechanisms that give rise to multiple drug recognition in these efflux transporters and their regulators. We have determined several x-ray structures of these important proteins, including the E. coli AcrB efflux transporter, C. jejuni CmeR transcriptional regulator and E. coli CusB heavy-metal efflux protein. We also crystallized the N. gonorrhoeae NorM multidrug transporter and collected the x-ray diffraction data of the crystals. To gain further insight into the mechanism of multiple drug recognition, we examined the binding affinities of AcrB and AcrR to different drugs using fluorescence polarization assays. In this thesis, we will summarize the new findings with AcrB, AcrR, CmeR and CusB, and discuss the structure and function of these efflux transporters and regulators

Ensuring Teaching Continuity: Chilean University Students’ Perception on Remote Teaching of English during COVID 19 Pandemic

The objective of our study is to know the perception of Chilean university students toward the pedagogical methodologies in online English classes during the COVID-19 pandemic. We applied a questionnaire (built by fifteen experts, validated through a pilot test of 34 students, and improved with licensed 21.0 version SPSS in terms of consistency) to 341 students from different university programs. We analyzed the responses quantitatively and qualitatively and found out that 75% of the 341 students surveyed have shown agreement with the pedagogical methodologies applied in online classes and motivation in learning due to the permanent concern and communication of the teacher in charge. Besides, the students have also yielded satisfactory learning outcomes. These results reveal that the online English teaching of Chilean higher education during the pandemic has been good and indirectly show the willingness of Chilean university teachers to receive relevant and continuous training to deliver quality education. In 2022, many countries have returned to face-to-face classes, but we believe that the pedagogical methodologies used in this research could serve as a reference for teachers or researchers worldwide who wish to incorporate into their face-to-face classes a part of online teaching and do research on online English teaching

Radiographic Disclosure of Fournier’s Gangrene

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Molecular and clinical analyses of 84 patients with tuberous sclerosis complex

BACKGROUND: Tuberous sclerosis complex (TSC) is an autosomal dominant disease characterized by the development of multiple hamartomas in many internal organs. Mutations in either one of 2 genes, TSC1 and TSC2, have been attributed to the development of TSC. More than two-thirds of TSC patients are sporadic cases, and a wide variety of mutations in the coding region of the TSC1 and TSC2 genes have been reported. METHODS: Mutational analysis of TSC1 and TSC2 genes was performed in 84 Taiwanese TSC families using denaturing high-performance liquid chromatography (DHPLC) and direct sequencing. RESULTS: Mutations were identified in a total of 64 (76 %) cases, including 9 TSC1 mutations (7 sporadic and 2 familial cases) and 55 TSC2 mutations (47 sporadic and 8 familial cases). Thirty-one of the 64 mutations found have not been described previously. The phenotype association is consistent with findings from other large studies, showing that disease resulting from mutations to TSC1 is less severe than disease due to TSC2 mutation. CONCLUSION: This study provides a representative picture of the distribution of mutations of the TSC1 and TSC2 genes in clinically ascertained TSC cases in the Taiwanese population. Although nearly half of the mutations identified were novel, the kinds and distribution of mutation were not different in this population compared to that seen in larger European and American studies

Traditional Chinese Medicine Diagnosis “ Yang-Xu Zheng

Pathogenesis of sepsis includes complex interaction between pathogen activities and host response, manifesting highly variable signs and symptoms, possibly delaying diagnosis and timely life-saving interventions. This study applies traditional Chinese medicine (TCM) Zheng diagnosis in patients with severe sepsis and septic shock to evaluate its adaptability and use as an early predictor of sepsis mortality. Three-year prospective observational study enrolled 126 septic patients. TCM Zheng diagnosis, Acute Physiology and Chronic Health Evaluation (APACHE) II score, and blood samples for host response cytokines measurement (tumor necrosis factor-α, Interleukin-6, Interleukin-8, Interleukin-10, Interleukin-18) were collected within 24 hours after admission to Intensive Care Unit. Main outcome was 28-day mortality; multivariate logistic regression analysis served to determine predictive variables of the sepsis mortality. APACHE II score, frequency of Nutrient-phase heat, and Qi-Xu and Yang-Xu Zhengs were significantly higher in nonsurvivors. The multivariate logistic regression analysis identified Yang-Xu Zheng as the outcome predictor. APACHE II score and levels of five host response cytokines between patients with and without Yang-Xu Zheng revealed significant differences. Furthermore, cool extremities and weak pulse, both diagnostic signs of Yang-Xu Zheng, were also proven independent predictors of sepsis mortality. TCM diagnosis “Yang-Xu Zheng” may provide a new mortality predictor for septic patients

Conformational change of the AcrR regulator reveals a possible mechanism of induction

The Escherichia coli AcrR multidrug-binding protein represses transcription of acrAB and is induced by many structurally unrelated cytotoxic compounds. The crystal structure of AcrR in space group P2221 has been reported previously. This P2221 structure has provided direct information about the multidrug-binding site and important residues for drug recognition. Here, a crystal structure of this regulator in space group P31 is presented. Comparison of the two AcrR structures reveals possible mechanisms of ligand binding and AcrR regulation

Comparison between the recovery time of alfentanil and fentanyl in balanced propofol sedation for gastrointestinal and colonoscopy: a prospective, randomized study

BACKGROUND: There is increasing interest in balanced propofol sedation (BPS) titrated to moderate sedation (conscious sedation) for endoscopic procedures. However, few controlled studies on BPS targeted to deep sedation for diagnostic endoscopy were found. Alfentanil, a rapid and short-acting synthetic analog of fentanyl, appears to offer clinically significant advantages over fentanyl during outpatient anesthesia. It is reasonable to hypothesize that low dose of alfentanil used in BPS might also result in more rapid recovery as compared with fentanyl. METHODS: A prospective, randomized and double-blinded clinical trial of alfentanil, midazolam and propofol versus fentanyl, midazolam and propofol in 272 outpatients undergoing diagnostic esophagogastroduodenal endoscopy (EGD) and colonoscopy for health examination were enrolled. Randomization was achieved by using the computer-generated random sequence. Each combination regimen was titrated to deep sedation. The recovery time, patient satisfaction, safety and the efficacy and cost benefit between groups were compared. RESULTS: 260 participants were analyzed, 129 in alfentanil group and 131 in fentanyl group. There is no significant difference in sex, age, body weight, BMI and ASA distribution between two groups. Also, there is no significant difference in recovery time, satisfaction score from patients, propofol consumption, awake time from sedation, and sedation-related cardiopulmonary complications between two groups. Though deep sedation was targeted, all cardiopulmonary complications were minor and transient (10.8%, 28/260). No serious adverse events including the use of flumazenil, assisted ventilation, permanent injury or death, and temporary or permanent interruption of procedure were found in both groups. However, fentanyl is New Taiwan Dollar (NT$) 103 (approximate US$ 4) cheaper than alfentanil, leading to a significant difference in total cost between two groups. CONCLUSIONS: This randomized, double-blinded clinical trial showed that there is no significant difference in the recovery time, satisfaction score from patients, propofol consumption, awake time from sedation, and sedation-related cardiopulmonary complications between the two most common sedation regimens for EGD and colonoscopy in our hospital. However, fentanyl is NT$103 (US$ 4) cheaper than alfentanil in each case. TRIAL REGISTRATION: Institutional Review Board of Buddhist Tzu Chi General Hospital (IRB097-18) and Chinese Clinical Trial Registry (ChiCTR-TRC-12002575

Peripheral Sympathectomy for Raynaud's Phenomenon: A Salvage Procedure

We retrospectively reviewed the effectiveness of peripheral sympathectomy for severe Raynaud's phenomenon. In this study, a total of 14 digits from six patients with chronic digital ischemic change were included. All patients had pain, ulcer, or gangrenous change in the affected digits and were unresponsive to pharmacologic or other nonsurgical therapies. In all cases, angiography showed multifocal arterial lesions, so microvascular reconstruction was unfeasible. Peripheral sympathectomy was performed as a salvage procedure to prevent digit amputation. The results were analyzed according to reduction of pain, healing of ulcers, and prevention of amputation. In 12 of the 14 digits, the ulcers healed and amputation was avoided. In the other two digits, the ulcers improved and progressive gangrene was limited. As a salvage procedure for Raynaud's phenomenon recalcitrant to conservative treatment, peripheral sympathectomy improves perfusion to ischemic digits and enables amputation to be avoided