Perceived stress and associated factors among medical students

Background: Stress and its psychological manifestations are currently a major source of concern. Medical education poses challenging and potentially threatening demands for students throughout the world. Objectives: To determine the prevalence and factors associated with perceived stress in medical students in the College of Medicine, King Saud Bin Abdulaziz University for Health Sciences, King Fahad Medical City, Riyadh, Saudi Arabia. Materials and Methods: This was a cross-sectional study on all medical students of batches 9, 10, and 11, which constituted all the enrolled students. Data were collected using a questionnaire based on the Kessler10 psychological distress instrument with a total score ranging from 10 to 50 points in addition to some sociodemographic characteristics. Appropriate statistical test procedures were used to study the magnitude of stress and its risk factors. Results: Mean stress score of the eighty participants was 26.03 ± 9.7. Students with severe stress constituted 33.8%, and 30% were well. Severe stress was significantly associated with female gender and junior level. Nervousness, feeling hopeless, feeling restless, and depressed were the most important factors affecting students′ stress scores. Factor analysis revealed three hidden factors for stress in this group, namely, depression, nervousness, and age. Conclusion: Stress in medical students is prevalent and significantly associated with the female gender and the junior level. Implementation of coping programs is necessary

Immunogenicity without Efficacy of an Adenoviral Tuberculosis Vaccine in a Stringent Mouse Model for Immunotherapy during Treatment

<div><p>To investigate if bacterial persistence during TB drug treatment could be overcome by modulation of host immunity, we adapted a clinically-relevant model developed for the evaluation of new drugs and examined if immunotherapy with two adenoviral vaccines, Ad35-TBS (AERAS-402) and Ad26-TBS, could shorten therapy in mice. Even though immunotherapy resulted in strong splenic IFN-γ responses, no effect on bacterial replication in the lungs was seen. Multiplex assay analysis of lung samples revealed the absence of cytokine augmentation such as IFN-γ, TNF-α and IL-2, suggesting that immunization failed to induce immunity in the lungs. In this model, we show that IFN-γ levels were not associated with protection against disease relapse. The results obtained from our study raise questions regarding the traits of protective TB immunity that are relevant for the development of future immunotherapeutic and post-exposure vaccination strategies.</p></div

Comparison of T-cell responses between animals receiving immunotherapy and chronically infected untreated animals.

<p>ELISpot assay was conducted to measure IFN-γ production in the TV Ad35+ and NC-IU animals following stimulation with A) CD4 peptide antigen to Ag85A B) CD8 peptide antigens to Ag85A and C) CD8 peptide antigens to Ag85B. Mice in the TV Ad35+ group received immunotherapy via intramuscular injection with 10¹⁰ viral particles of Ad35-TBS at weeks 4 and 8. Lines depict geometric mean and p-values were calculated using ANOVA followed by Tukey’s multiple comparisons test (n = 5 at each time point for all groups) and are detailed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0127907#pone.0127907.t002" target="_blank">Table 2</a>. SFU = splenocyte forming units.</p

Schematic of study design.

<p>420 mice were divided into animals that were uninfected (NC-UU) (20 mice) and those that were infected with <i>M</i>.<i>tb</i> at week -2 (NC-IU) (400 mice). At week 0, 365 mice from NC-IU were removed to create the PC cohort that was initiated on therapy with R, H and Z via oral gavage, 7 days per week. The remaining NC-IU animals were left untreated up to week 12, when they became moribund and were sacrificed. At week 4, 284 animals from the PC cohort were randomized into groups NC-T4, NC-V and TV Ad35+. At this time point, mice in the NC-V and TV Ad35+ groups received intramuscular immunotherapy with 10¹⁰ viral particles of Ad35 empty vector or Ad35-TBS vaccine, which were respectively boosted at week 8. Animals in the PC, NC-T4, NC-V and TV Ad35+ groups were switched to therapy with R and H at week 8 which lasted to week 16, except for the PC group which received therapy up to week 24. At week 16, when therapy was shortened, 56 mice from the TV Ad35+ group were removed and divided into 2 further groups, TV Ad26+ and TV Ad26- which received immunotherapy with Ad26-TBS and Ad26, respectively. All animals were followed through for 3 months following therapy cessation at week 16 and week 24 (PC). Animals were subsequently sacrificed for the final lung CFU enumeration. Multiplex analysis on lung samples were carried out at times indicated for CFU counting, from week 4 onwards.</p

Numbers of mice used in the study at each time point.

<p>420 female BALBc mice were used to obtain data for immunological analysis, relapse CFU measurements and histopathology. Numbers were gradually increased to ensure sufficient power to detect differences between the groups as described previously. NC-UU = negative control, uninfected, untreated; NC-IU = negative control, infected, untreated; PC = positive control, standard therapy; NC-T4 = negative control, shortened therapy; NC-V = negative control, Ad35 empty vector, shortened therapy; TV Ad35+ = Ad35-TBS immunotherapy, shortened therapy; TV Ad26+ = Ad26-TBS immunotherapy, shortened therapy; TV Ad26- = negative control, Ad26 empty vector, shortened therapy.</p><p>Numbers of mice used in the study at each time point.</p

Multiplex enzyme-linked immunosorbent assay of four key cytokines.

<p>At each time point 4 weeks post-treatment initiation where lungs were harvested for CFU counting, 150μl of lung homogenate was set aside for cytokine analysis. At week 4, animals in the TV Ad35+, NC-V, NC-T4 and PC groups were identical prior to randomization. Immediately following randomization at week 4, animals in the TV Ad35+ group received immunotherapy with 10¹⁰ viral particles of Ad35-TBS and animals in the NC-V group received 10¹⁰ empty Ad35 vector which were both boosted at week 8. At week 16, animals in the TV Ad35+ groups were randomized into TV Ad26+ and TV Ad26- and received 10¹⁰ viral particles of Ad26-TBS and Ad26 empty vector, respectively. All animals were administered anti-TB therapy except the NC-IU and NC-UU control groups as depicted in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0127907#pone.0127907.g001" target="_blank">Fig 1</a>. Graph depicts the mean cytokine value with standard deviation (n = 4 at each time point for all groups).</p

Immunogenicity of Ad35-TBS and Ad26-TBS immunotherapy.

<p>At week 4, baseline IFN-γ production to pooled peptides covering the whole sequence of Ag85A, Ag85B and TB10.4 were measured by ELISpot prior to randomization (refer to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0127907#pone.0127907.g001" target="_blank">Fig 1</a>). At this time point, the values between the NC-T4 and TV Ad35+ are identical. Following randomization, animals in the TV Ad35+ group received immunotherapy at weeks 4 and 8 with 10¹⁰ viral particles of Ad35-TBS intramuscularly. At week 16, baseline immune responses were measured in the TV Ad35+ group prior to randomization into TV Ad26+ and TV Ad26- groups. The TV Ad26+ animals then received heterologous boosting with 10¹⁰ viral particles of Ad26-TBS at week 16 whilst the TV Ad26- received empty Ad26 vector as control. p-values were calculated two weeks after each immunotherapy (weeks 6 and 10) and are detailed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0127907#pone.0127907.t002" target="_blank">Table 2</a> (ANOVA followed by Tukey’s multiple comparisons test, n = 5 at each time point for all groups). Lines depict geometric mean. No significant differences were observed following Ad26-TBS boosting at week 16 for all groups (NS = not significant, t-test). SFU = splenocyte forming units.</p

T-cell responses following Ad35-TBS and Ad26-TBS immunotherapy.

<p>At week 4, baseline IFN-γ production following stimulation with CD4 and CD8 peptides to Ag85A and CD8 peptides to Ag85B were measured by ELISpot prior to randomization (refer to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0127907#pone.0127907.g001" target="_blank">Fig 1</a>). At week 4, animals in the NC-T4 and TV Ad35+ have not been randomized and are identical to one another. Immediately following randomization at week 4 into the respective groups, immunotherapy was administered via intramuscular injection to the TV Ad35+ animals with 10¹⁰ viral particles of Ad35-TBS which was boosted at week 8. At week 16, baseline immune responses were measured in the TV Ad35+ group prior to randomization into TV Ad26+ and TV Ad26- groups. The TV Ad26+ animals then received heterologous boosting with 10¹⁰ viral particles of Ad26-TBS at week 16 whilst the TV Ad26- received empty Ad26 vector as control. p-values were calculated two weeks after each immunotherapy (weeks 6 and 10) and are detailed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0127907#pone.0127907.t002" target="_blank">Table 2</a> (n = 5 at each time point for all groups). Lines depict geometric mean. Boosting with Ad26-TBS resulted in significant heightening of CD8 responses to Ag85A (p = 0.01, t-test) and Ag85B (p = 0.01, t-test) at week 18 compared to week 16 that was not observed for CD4 responses to Ag85A (NS = not significant, p = 0.66, t-test). SFU = splenocyte forming units.</p

Probing the mechanism of failure at week 28.

<p>A) ELISpot analysis was conducted at the final relapse time point of week 28 as previously described. Results of groups that received immunotherapy (TV Ad35+ and TV Ad26+) and the main negative control group NC-T4 are depicted. Results of significance testing are depicted by * for comparison between TV Ad35+ and NC-T4 whereby: * p<0.05, ** p<0.01, *** p<0.001 and **** p<0.0001. § denotes significant differences between TV Ad26+ and NC-T4 whereby: § p<0.05, §§ p<0.01, §§§ p<0.001 and §§§§ p<0.0001. Statistical significance was determined by ANOVA followed by Tukey’s multiple comparisons test where n = 5. B) Multiplex enzyme-linked immunosorbent assay was carried out at week 28 on lung samples as previously described. Results from groups that received immunotherapy (TV Ad35+ and TV Ad26+) and the main control group NC-T4 are shown (mean and standard deviation, n = 4 per group). No statistically significant differences in the levels of IFN-γ, TNF-α or IL-2 were measured between all groups (IFN-γ: p = 0.35; TNF-α: p = 0.98; IL-2: p = 0.09; ANOVA). C) Histopathology of lung samples at week 12 was examined for signs of immunopathology following two doses of immunotherapy with Ad35-TBS. In comparison to the NC-IU animals, the lungs of animals that underwent therapy appeared similar with minimal signs of lesions, necrosis or significant immune infiltration. Immunotherapy did not result in observable immunopathology. Results of histopathology at week 12 are representative of all time points measured (weeks 4, 6, 8 and 10) and at week 18 following Ad26-TBS boosting (data not shown).</p