Listeria monocytogenes induces host DNA damage and delays the host cell cycle to promote infection

Listeria monocytogenes (Lm) is a human intracellular pathogen widely used to uncover the mechanisms evolved by pathogens to establish infection. However, its capacity to perturb the host cell cycle was never reported. We show that Lm infection affects the host cell cycle progression, increasing its overall duration but allowing consecutive rounds of division. A complete Lm infectious cycle induces a S-phase delay accompanied by a slower rate of DNA synthesis and increased levels of host DNA strand breaks. Additionally, DNA damage/replication checkpoint responses are triggered in an Lm dose-dependent manner through the phosphorylation of DNA-PK, H2A.X, and CDC25A and independently from ATM/ATR. While host DNA damage induced exogenously favors Lm dissemination, the override of checkpoint pathways limits infection. We propose that host DNA replication disturbed by Lm infection culminates in DNA strand breaks, triggering DNA damage/replication responses, and ensuring a cell cycle delay that favors Lm propagation.     

GRASP55 regulates the unconventional secretion and aggregation of mutant huntingtin

Recent studies demonstrated that the Golgi reassembly stacking proteins (GRASPs), especially GRASP55, regulate Golgi-independent unconventional secretion of certain cytosolic and transmembrane cargoes; however, the underlying mechanism remains unknown. Here, we surveyed several neurodegenerative disease–related proteins, including mutant huntingtin (Htt-Q74), superoxide dismutase 1 (SOD1), tau, and TAR DNA–binding protein 43 (TDP-43), for unconventional secretion; our results show that Htt-Q74 is most robustly secreted in a GRASP55-dependent manner. Using Htt-Q74 as a model system, we demonstrate that unconventional secretion of Htt is GRASP55 and autophagy dependent and is enhanced under stress conditions such as starvation and endoplasmic reticulum stress. Mechanistically, we show that GRASP55 facilitates Htt secretion by tethering autophagosomes to lysosomes to promote autophagosome maturation and subsequent lysosome secretion and by stabilizing p23/TMED10, a channel for translocation of cytoplasmic proteins into the lumen of the endoplasmic reticulum–Golgi intermediate compartment. Moreover, we found that GRASP55 levels are upregulated by various stresses to facilitate unconventional secretion, whereas inhibition of Htt-Q74 secretion by GRASP55 KO enhances Htt aggregation and toxicity. Finally, comprehensive secretomic analysis identified novel cytosolic cargoes secreted by the same unconventional pathway, including transgelin (TAGLN), multifunctional protein ADE2 (PAICS), and peroxiredoxin-1 (PRDX1). In conclusion, this study defines the pathway of GRASP55-mediated unconventional protein secretion and provides important insights into the progression of Huntington’s disease.     

FAS/FASL Expression Profile as a Prognostic Marker in Squamous Cell Carcinoma of the Oral Cavity

FAS/FASL altered expression may cause tumor protecting immunomodulation, with a direct impact on patient prognosis. FAS expression was studied in 60 squamous cell carcinomas of the oral cavity. FAS expression did not show a significant association with tumor histopathological characteristics, but was significantly associated with lymph node positivity. FAS expression was significantly associated with disease specific death and negative FAS expression was an independent risk factor, increasing risk 4 times when compared to positive expression. When FAS and FASL expression results were combined, we were able to define high, intermediate and low risk profiles. Disease-free and disease-specific survival were significantly correlated with FAS/FASL expression profiles. The high risk category was an independent marker for earlier disease relapse and disease-specific death, with approximately 4- and 6-fold increased risk, respectively, when compared to the low risk profile. Risk profiles based on FAS/FASL expression showed that high risk was significantly associated with increased disease relapse and death, as well as shorter disease-free or disease-specific survival. This categorization, added to patient clinical data, may facilitate the choice of therapy, minimizing treatment failure and increasing disease control.     

Effect of genetic modifications by selection for immunological tolerance on fungus infection in mice

Two strains of mice genetically selected for extreme phenotypes of immunological tolerance to ovalbumin, susceptible (TS) and resistant (TR), were experimentally infected with Sporothrix schenckii. The objective was to observe whether the genetic modifications produced by the selection might be associated with interstrain differences in adaptive immune and innate responses to infection. Therefore, we evaluated the LD(50), CFU, phagocytic index, fungicidal activity, pro-inflammatory cytokines, specific antibody titres, and the delayed-type hypersensitivity reactivity. TR mice were tenfold more susceptible to infection than TS mice, as shown by LD(50) (5 x 10(6) conidia i.v.). In TS mice, the resistance was a consequence of the tissue fungal load reduction, consistent specific T-cell-mediated immunity, and tumour necrosis factor (TNF)-alpha activity at onset of infection. In TR mice, these responses were not precociously detected. Therefore, the absence of CD4(+) T-cell response in the first week of infection might explain the non-clearance of pathogen in TR mice. However, TR mice did show an increase in TNF level and delayed-type hypersensitivity response after the first week post-infection; there was also expansion and increase in granulomatous foci and CFU in the spleen. The expansion of granulomatous foci and the increase in TNF-alpha and tissue fungal load to damaging levels induced severe tissue destruction, general failure of the organs, cachexy and death in TR mice. The results show that genetic selection for extreme phenotypes of immunological tolerance also modified the responses to S. schenckii infection.     

T Cell Activation and Cytokine Profile of Tuberculosis and HIV-Positive Individuals during Antituberculous Treatment and Efavirenz-Based Regimens

Introduction

The profile of immune activation markers in tuberculosis and HIV-infected patients is already known. The impact of simultaneous infections on the immune parameters is still not fully explored.

Methods

We conducted a prospective study to estimate trajectories of activated T cell subsets and the profile of anti- and pro-inflammatory cytokines in a group of HIV-TB individuals, previously naïve for HAART, recruited from a randomized clinical trial during TB treatment and first antiretroviral therapy with efavirenz. Patients were evaluated according to the immunosuppression levels at baseline as group 1 (CD4<200 cells/mm³) and group 2 (CD4>200 cells/mm³). These parameters were measured at the time of HAART initiation (started about 30 days after the onset of TB treatment) and at the follow-up visits after 30, 60, 90 and 180 days. Trajectories were estimated using least squares estimates of the coefficients of a restricted cubic spline function in time after adjusting for subject effects, bootstrapping it 500 times.

Results

Increase of CD4 T cell counts and suppression of HIV viral load were observed for all patients under HAART and TB treatment. Descendent trajectories were observed for the activated CD8⁺/CD38⁺ and CD3⁺/HLA-DR⁺ T cell subsets, and for plasma concentration of gamma- interferon (IFN-γ). Except for TNF-α and IL-2 discrete variations were observed for the other cytokines. Differences in the trajectories of these parameters were observed for groups 1 and 2. Higher values of IFN-γ, IL-2, IL-6 and IL-10 were observed for group 1 from the baseline to two months after treatment initiation, whereas reduced levels of TNF-α were observed for this group between 60 and 120 days of HAART.

Conclusion

Independent of the immunosuppression profile at baseline, HIV-TB patients under HAART were able to recover the CD4⁺ T cell counts, and control viral replication and immune activation parameters over time.     

Parasitism in Trialeurodes variabilis (Quaintance) (Hemiptera: Aleyrodidae) by Encarsia hispida De Santis (Hymenoptera: Aphelinidae), in papaya, in Brazil

Infestation of Trialeurodes variabilis (Quaintance) was observed in October 2004, in papaya plants of cultivar Sunrise Solo, under screenhouse conditions, in Cruz das Almas, State of Bahia, Brazil. In infested leaves, around 20% of parasitism on nymphs was verified. Leaves with parasitized nymphs were kept in laboratory until emergence of the parasitoid, identified as Encarsia hispida De Santis. This is the first time that this parasitoid was detected on T. variabilis nymphs in Brazil.     

Cervicovaginal cytology in uterine adenocarcinoma and adenosquamous carcinoma. Comparison of cytologic and histologic findings

To investigate the diagnostic accuracy and to characterize the findings in false-negative cases, the results of cervicovaginal cytology in 56 adenocarcinomas and 25 adenosquamous carcinomas (42 cervical, 36 endometrial, 2 metastatic and 1 arising synchronously from both cervix and endometrium) were reviewed, including review of the actual slides in 56 cases. Overall, 80% of the initial cytologic diagnoses resulted in diagnostic curettage (i.e., cytology was effectively positive); 84% of the postreview diagnosis were effectively positive. Nine cytology slides showed no malignant cells; eight of these negative smears showed repair, five were atrophic, two showed a high estrogen effect and one had enlarged atypical bare nuclei. These false-negative diagnoses were associated with an endometrial primary site (P less than .01), endometrioid histology (P less than .005), low-grade or intermediate-grade histology (P less than .005), small size of tumor (P less than .05) and absence of cervical involvement (P less than .005) in those cases in which a hysterectomy was performed. False-negative diagnoses were not associated with an absence of endocervical cells or with scanty cellularity. Of 39 cervical and 28 endometrial carcinomas with a positive cytologic diagnosis (initially or after review of the available slides), cytology correctly identified the primary site in 18% and 54% of the cases, respectively. Cytology incorrectly classified the anatomic site of four cervical and three endometrial carcinomas and considered one case arising in both the endometrium and cervix to be endometrial. Routine cervicovaginal cytology does have a role in screening for uterine glandular carcinoma; to maximize its diagnostic sensitivity, we suggest using a recommendation for curettage in the report of positive cases so that all of the varied cytologic diagnoses associated with glandular carcinomas will receive a uniform clinical response. In those cases with preserved cancer cells, a correlation can be made with the histologic type of the carcinoma, rather than with the anatomic site.