The role of integrins in Drosophila egg chamber morphogenesis

A Drosophila egg chamber is comprised of a germline cyst surrounded by a tightly-organised epithelial monolayer, the follicular epithelium (FE). Loss of integrin function from the FE disrupts epithelial organisation at egg chamber termini, but the cause of this phenotype remains unclear. Here we show that the -integrin Myospheroid (Mys) is only required during early oogenesis when the pre-follicle cells form the FE. mys mutants disrupt both the formation of a monolayered epithelium at egg chamber termini and the morphogenesis of the stalk between adjacent egg chambers, which develops through the intercalation of two rows of cells into a single-cell wide stalk. Secondary epithelia, like the FE, have been proposed to require adhesion to the basement membrane to polarise. However, Mys is not required for pre-follicle cell polarisation, as both follicle and stalk cells localise polarity factors correctly, despite being mispositioned. Instead, loss of integrins causes pre-follicle cells to basally constrict, detach from the basement membrane and become internalised. Thus, integrin function is dispensable for pre-follicle cell polarity but is required to maintain cellular organisation and cell shape during morphogenesis.This work was supported by a Wellcome Trust Principal Fellowship to D.St J. (080007, 207496) and by centre grant support from the Wellcome Trust (092096, 203144) and Cancer Research UK (A14492, A24823). H.E.L. was supported by a Herchel Smith Studentship, University of Cambridge. D.T.B. was supported by a Marie Curie Fellowship (funded by the European Commission) and the Wellcome Trust

Pins is not required for spindle orientation in the Drosophila wing disc.

In animal cells, mitotic spindles are oriented by the dynein/dynactin motor complex, which exerts a pulling force on astral microtubules. Dynein/dynactin localization depends on Mud/NUMA, which is typically recruited to the cortex by Pins/LGN. In Drosophila neuroblasts, the Inscuteable/Baz/Par-6/aPKC complex recruits Pins apically to induce vertical spindle orientation, whereas in epithelial cells Dlg recruits Pins laterally to orient the spindle horizontally. Here we investigate division orientation in the Drosophila imaginal wing disc epithelium. Live imaging reveals that spindle angles vary widely during prometaphase and metaphase, and therefore do not reliably predict division orientation. This finding prompted us to re-examine mutants that have been reported to disrupt division orientation in this tissue. Loss of Mud misorients divisions, but Inscuteable expression and aPKC, dlg and pins mutants have no effect. Furthermore, Mud localizes to the apical-lateral cortex of the wing epithelium independently of both Pins and cell cycle stage. Thus, Pins is not required in the wing disc because there are parallel mechanisms for Mud localization and hence spindle orientation, making it a more robust system than in other epithelia.This work was supported by a Wellcome Trust Principal Fellowship to DStJ [080007] and by core support from the Wellcome Trust [092096] and Cancer Research UK [A14492]. DTB was supported by a Marie Curie Fellowship and the Wellcome Trust. HEL was supported by a Herchel Smith Studentship.This is the author accepted manuscript. The final version is available from The Company of Biologists via http://dx.doi.org/10.1242/dev.13547

Lateral adhesion drives reintegration of misplaced cells into epithelial monolayers.

Cells in simple epithelia orient their mitotic spindles in the plane of the epithelium so that both daughter cells are born within the epithelial sheet. This is assumed to be important to maintain epithelial integrity and prevent hyperplasia, because misaligned divisions give rise to cells outside the epithelium. Here we test this assumption in three types of Drosophila epithelium; the cuboidal follicle epithelium, the columnar early embryonic ectoderm, and the pseudostratified neuroepithelium. Ectopic expression of Inscuteable in these tissues reorients mitotic spindles, resulting in one daughter cell being born outside the epithelial layer. Live imaging reveals that these misplaced cells reintegrate into the tissue. Reducing the levels of the lateral homophilic adhesion molecules Neuroglian or Fasciclin 2 disrupts reintegration, giving rise to extra-epithelial cells, whereas disruption of adherens junctions has no effect. Thus, the reinsertion of misplaced cells seems to be driven by lateral adhesion, which pulls cells born outside the epithelial layer back into it. Our findings reveal a robust mechanism that protects epithelia against the consequences of misoriented divisions.The authors are grateful to R. Nieuwburg, the St Johnston group, and other Gurdon Institute members for suggestions. We thank the Bloomington Stock Center, J. Knoblich, and the TRiP at Harvard Medical School (NIH/NIGMS R01-GM084947) for fly stocks. We thank N. Lowe for technical assistance. This work was supported by a Wellcome Trust Principal Fellowship to D.St.J. (080007), and by core support from the Wellcome Trust (092096) and Cancer Research UK (A14492). D.T.B. was supported by a Marie Curie Fellowship and the Wellcome Trust. H.E.L. was supported by a Herchel Smith Studentship.This is the author accepted manuscript. The final version is available from NPG via http://dx.doi.org/10.1038/ncb324

Outcomes of Multi-Drug Resistant Tuberculosis (MDR-TB) among a Cohort of South African Patients with High HIV Prevalence

Multidrug-resistant tuberculosis (MDR-TB) is a major clinical challenge, particularly in patients with human immunodeficiency virus (HIV) co-infection. MDR-TB treatment is increasingly available, but outcomes have not been well characterized. South Africa has provided MDR-TB treatment for a decade, and we evaluated outcomes by HIV status for patients enrolled between 2000 and 2004 prior to anti-retroviral access.We assessed treatment outcomes in a prospective cohort of patients with MDR-TB from eight provincial programs providing second line drugs. World Health Organization definitions were used. Results were stratified by HIV status.Seven hundred fifty seven patients with known HIV status were included in the final analysis, and HIV infection was documented in 287 (38%). Overall, 348 patients (46.0%) were successfully treated, 74 (9.8%) failed therapy, 177 (23.4%) died and 158 (20.9%) defaulted. Patients with HIV were slightly younger and less likely to be male compared to HIV negative patients. Patients with HIV were less likely to have a successful treatment outcome (40.0 vs. 49.6; P<0.05) and more likely to die (35.2 vs. 16.2; P<0.0001). In a competing risk survival analysis, patients with HIV had a higher hazard of death (HR: 2.33, P<0.0001). Low baseline weight (less than 45 kg and less than 60 kg) was also associated with a higher hazard of death (HR: 2.52, P<0.0001; and HR: 1.50, P<0.0001, respectively, compared to weight greater than 60 kg). Weight less than 45 kg had higher risk of failure (HR: 3.58, P<0.01). Any change in treatment regimen was associated with a higher hazard of default (HR: 2.86; 95% CI 1.55-5.29, P<0.001) and a lower hazard of death (HR: 0.63, P<0.05).In this MDR-TB treatment program patients with HIV infection and low weight had higher hazards of death. Overall treatment outcomes were poor. Efforts to improve treatment for MDR-TB are urgently needed

Neisseria gonorrhoeae Activates the Proteinase Cathepsin B to Mediate the Signaling Activities of the NLRP3 and ASC-Containing Inflammasome

Neisseria gonorrhoeae is a common sexually transmitted pathogen that significantly impacts female fertility, neonatal health, and transmission of HIV worldwide. N. gonorrhoeae usually causes localized inflammation of the urethra and cervix by inducing production of IL-1β and other inflammatory cytokines. Several NLR (Nucleotide binding domain, Leucine Rich Repeat) proteins are implicated in the formation of pro-IL-1β-processing complexes called inflammasomes in response to pathogens. We demonstrate that NLRP3 (cryopyrin,NALP3) is the primary NLR required for IL-1β/IL-18 secretion in response to N. gonorrhoeae in monocytes. We also show that N. gonorrhoeae infection promotes NLRP3-dependent monocytic cell death via pyronecrosis, a recently described pathway with morphological features of necrosis, including release of the strong inflammatory mediator HMBG1. Additionally, N. gonorrhoeae activates the cysteine protease Cathepsin-B as measured by the breakdown of a Cathepsin B substrate. Inhibition of Cathepsin B shows that this protease is an apical controlling step in the downstream activities of NLRP3 including IL-1β production, pyronecrosis, and HMGB1 release. Non-pathogenic Neisseria strains (N. cinerea and N. flavescens) do not activate NLRP3 as robustly as N. gonorrhoeae. Conditioned media from N. gonorrhoeae contains factors capable of initiating the NLRP3 mediated signaling events. Isolated N. gonorrhoeae lipooligosaccharide, a known virulence factor from this bacterium that is elaborated from the bacterium in the form of outer membrane blebs, activates both NLRP3-induced IL-1β secretion and pyronecrosis. Our findings indicate that activation of NLRP3-mediated inflammatory response pathways is an important venue associated with host response and pathogenesis of N. gonorrhoeae

A Tissue Biomarker Panel Predicting Systemic Progression after PSA Recurrence Post-Definitive Prostate Cancer Therapy

Many men develop a rising PSA after initial therapy for prostate cancer. While some of these men will develop a local or metastatic recurrence that warrants further therapy, others will have no evidence of disease progression. We hypothesized that an expression biomarker panel can predict which men with a rising PSA would benefit from further therapy.A case-control design was used to test the association of gene expression with outcome. Systemic (SYS) progression cases were men post-prostatectomy who developed systemic progression within 5 years after PSA recurrence. PSA progression controls were matched men post-prostatectomy with PSA recurrence but no evidence of clinical progression within 5 years. Using expression arrays optimized for paraffin-embedded tissue RNA, 1021 cancer-related genes were evaluated-including 570 genes implicated in prostate cancer progression. Genes from 8 previously reported marker panels were included. A systemic progression model containing 17 genes was developed. This model generated an AUC of 0.88 (95% CI: 0.84-0.92). Similar AUCs were generated using 3 previously reported panels. In secondary analyses, the model predicted the endpoints of prostate cancer death (in SYS cases) and systemic progression beyond 5 years (in PSA controls) with hazard ratios 2.5 and 4.7, respectively (log-rank p-values of 0.0007 and 0.0005). Genes mapped to 8q24 were significantly enriched in the model.Specific gene expression patterns are significantly associated with systemic progression after PSA recurrence. The measurement of gene expression pattern may be useful for determining which men may benefit from additional therapy after PSA recurrence

Association between vitamin D insufficiency and tuberculosis in a vietnamese population

<p>Abstract</p> <p>Background</p> <p>Recent <it>in vitro </it>evidence suggests a link between vitamin D status and the risk of tuberculosis (TB). This study sought to examine the association between vitamin D status, parathyroid hormone (PTH) and the risk of TB in a Vietnamese population.</p> <p>Methods</p> <p>The study was designed as a matched case-control study, which involved 166 TB patients (113 men and 53 women), who were age-and-sex matched with 219 controls (113 men and 106 women). The average age of men and women was 49 and 50, respectively. TB was diagnosed by the presence of acid-fast bacilli on smears from sputum, and the isolation of <it>M. tuberculosis</it>. All patients were hospitalized for treatment in a TB specialist hospital. Controls were randomly drawn from the general community within the Ho Chi Minh, Vietnam. 25-hydroxyvitamin D [25(OH)D] and PTH was measured prior to treatment by an electrochemiluminescence immunoassay (ECLIA) on a Roche Elecsys. A serum level of 25(OH)D below 30 ng/mL was deemed to be vitamin D insufficient.</p> <p>Results</p> <p>The prevalence of vitamin D insufficiency was 35.4% in men with TB and 19.5% in controls (<it>P </it>= 0.01). In women, there were no significant differences in serum 25(OH)D and serum PTH levels between TB patients and controls. The prevalence of vitamin D insufficiency in women with TB (45.3%) was not significantly different from those without TB (47.6%; <it>P </it>= 0.91). However, in both genders, serum calcium levels in TB patients were significantly lower than in non-TB individuals. Smoking (odds ratio [OR] 1.25; 95% confidence interval [CI] 1.10 - 14.7), reduced 25(OH)D (OR per standard deviation [SD]: 1.14; 95% CI 1.07 - 10.7) and increased PTH (OR per SD 1.13; 95% CI 1.05 - 10.4) were independently associated with increased risk of TB in men.</p> <p>Conclusion</p> <p>These results suggest that vitamin D insufficiency was a risk factor for tuberculosis in men, but not in women. However, it remains to be established whether the association is a causal relationship.</p