Perinatal Ureaplasma Exposure Is Associated With Increased Risk of Late Onset Sepsis and Imbalanced Inflammation in Preterm Infants and May Add to Lung Injury

Background: Controversy remains concerning the impact of Ureaplasma on preterm neonatal morbidity.Methods: Prospective single-center study in very low birth weight infants <30 weeks' gestation. Cord blood and initial nasopharyngeal swabs were screened for Ureaplasma parvum and U. urealyticum using culture technique and polymerase chain reaction. Neonatal outcomes were followed until death or discharge. Multi-analyte immunoassay provided cord blood levels of inflammatory markers. Using multivariate regression analyses, perinatal Ureaplasma exposure was evaluated as risk factor for the development of bronchopulmonary dysplasia (BPD), other neonatal morbidities until discharge and systemic inflammation at admission.Results: 40/103 (39%) infants were positive for Ureaplasma in one or both specimens, with U. parvum being the predominant species. While exposure to Ureaplasma alone was not associated with BPD, we found an increased risk of BPD in Ureaplasma-positive infants ventilated ≥5 days (OR 1.64; 95% CI 0.12–22.98; p = 0.009). Presence of Ureaplasma was associated with a 7-fold risk of late onset sepsis (LOS) (95% CI 1.80–27.39; p = 0.014). Moreover, Ureaplasma-positive infants had higher I/T ratios (b 0.39; 95% CI 0.08–0.71; p = 0.014), increased levels of interleukin (IL)-17 (b 0.16; 95% CI 0.02–0.30; p = 0.025) and matrix metalloproteinase 8 (b 0.77; 95% CI 0.10–1.44; p = 0.020), decreased levels of IL-10 (b −0.77; 95% CI −1.58 to −0.01; p = 0.043) and increased ratios of Tumor necrosis factor-α, IL-8, and IL-17 to anti-inflammatory IL-10 (p = 0.003, p = 0.012, p < 0.001).Conclusions: Positive Ureaplasma screening was not associated with BPD. However, exposure contributed to BPD in infants ventilated ≥5 days and conferred an increased risk of LOS and imbalanced inflammatory cytokine responses

A prospective study on histone γ-H2AX and 53BP1 foci expression in rectal carcinoma patients: correlation with radiation therapy-induced outcome

Background The prognostic value of histone γ-H2AX and 53BP1 proteins to predict the radiotherapy (RT) outcome of patients with rectal carcinoma (RC) was evaluated in a prospective study. High expression of the constitutive histone γ-H2AX is indicative of defective DNA repair pathway and/or genomic instability, whereas 53BP1 (p53-binding protein 1) is a conserved checkpoint protein with properties of a DNA double-strand breaks sensor. Methods Using fluorescence microscopy, we assessed spontaneous and radiation-induced foci of γ-H2AX and 53BP1 in peripheral blood mononuclear cells derived from unselected RC patients (n = 53) undergoing neoadjuvant chemo- and RT. Cells from apparently healthy donors (n = 12) served as references. Results The γ-H2AX assay of in vitro irradiated lymphocytes revealed significantly higher degree of DNA damage in the group of unselected RC patients with respect to the background, initial (0.5 Gy, 30 min) and residual (0.5 Gy and 2 Gy, 24 h post-radiation) damage compared to the control group. Likewise, the numbers of 53BP1 foci analyzed in the samples from 46 RC patients were significantly higher than in controls except for the background DNA damage. However, both markers were not able to predict tumor stage, gastrointestinal toxicity or tumor regression after curative RT. Interestingly, the mean baseline and induced DNA damage was found to be lower in the group of RC patients with tumor stage IV (n = 7) as compared with the stage III (n = 35). The difference, however, did not reach statistical significance, apparently, because of the limited number of patients. Conclusions The study shows higher expression of γ-H2AX and 53BP1 foci in rectal cancer patients compared with healthy individuals. Yet the data in vitro were not predictive in regard to the radiotherapy outcome

CIP2A Influences Survival in Colon Cancer and Is Critical for Maintaining Myc Expression

The cancerous inhibitor of protein phosphatase 2A (CIP2A) is an oncogenic factor that stabilises the c-Myc protein. CIP2A is overexpressed in several tumours, and expression levels are an independent marker for long-term outcome. To determine whether CIP2A expression is elevated in colon cancer and whether it might serve as a prognostic marker for survival, we analysed CIP2A mRNA expression by real-time PCR in 104 colon cancer samples. CIP2A mRNA was overexpressed in colon cancer samples and CIP2A expression levels correlated significantly with tumour stage. We found that CIP2A serves as an independent prognostic marker for disease-free and overall survival. Further, we investigated CIP2A-dependent effects on levels of c-Myc, Akt and on cell proliferation in three colon cancer cell lines by silencing CIP2A using small interfering (si) and short hairpin (sh) RNAs. Depletion of CIP2A substantially inhibited growth of colon cell lines and reduced c-Myc levels without affecting expression or function of the upstream regulatory kinase, Akt. Expression of CIP2A was found to be dependent on MAPK activity, linking elevated c-Myc expression to deregulated signal transduction in colon cancer

Fever-range temperature modulates activation and function of human dendritic cells stimulated with the pathogenic mould Aspergillus fumigatus

In immunocompromised patients, invasive aspergillosis (IA) is the most frequent disease caused by the pathogenic mould Aspergillus fumigatus. Fever is one of the most common yet nonspecific clinical symptoms of IA. To evaluate the role of hyperthermia in the innate immune response to A. fumigatus in vitro, human monocyte-derived dendritic cells (DCs) were stimulated with germ tubes of A. fumigatus or the fungal cell wall component zymosan at 37◦C or 40◦C, followed by characterization of specific DC functions. While maturation of DCs was enhanced and DC phagocytic capacity was reduced at 40◦C, we observed that DC viability and cytokine release were unaffected. Thus, our results suggest that hyperthermia has substantial impacts on DC function in vitro, which might also influence the course and outcome of IA in immunocompromised patients

Additional file 1: Table S1. of A prospective study on histone γ-H2AX and 53BP1 foci expression in rectal carcinoma patients: correlation with radiation therapy-induced outcome

Characteristics of healthy individuals and RC patients undergoing chemo-radiotherapy (Summary). Table S2. Patients’ characteristics in regard to chemo-radiation toxicities and alcohol/tobacco consumption. Table S3. DNA damage measured by the histone γ-H2AX in PBMCs isolated from blood of apparently healthy donors (N) and unselected rectal carcinoma (RC) patients after exposure to 0.5 or 2 Gy of X-irradiation in vitro or after 5 clinical radiation fractions. Table S4. DNA damage measured by the 53BP1 foci in PBMCs isolated from blood of apparently healthy donors (N) and unselected rectal carcinoma (RC) patients after exposure to 0.5 or 2 Gy of X-irradiation in vitro or after 5 clinical radiation fractions. Figure S1. DNA damage assessed by the mean number of 53BP1 foci in non-irradiated (A) and irradiated (B-D) PBMCs derived from unselected RC patients (triangles), as compared to cells from apparently healthy donors (circles). For further details, see legend to Fig. 1. Filled squares represent the mean values (± SE) for the respective group. “n.s.” indicates that the difference was not highly significant (p > 0.05). Figure S2. Correlational analysis of mean γ-H2AX and 53BP1 foci counts from 500 nuclei per sample. Non-irradiated (A) and irradiated with 0.5 (B and C) and 2 Gy (D) lymphocytes were fixed 30 min (B) or 24 h (C, D) post-IR. The expression of both proteins was analyzed simultaneously at each time and IR points for n = 48 blood samples derived from unselected RC patients. Figure S3. DNA damage assessed by means of the 53BP1 assay in non-irradiated (A) and irradiated (B-D) PBMCs derived from normally-reacting RC patients (grade 0 and 1, up triangles) and radiation-sensitive (grade 2 and 3, down triangles) cancer patients compared to cells from apparently healthy donors (circles). Filled squares represent the mean values (± SE) for the respective group. For details, see legend to Fig. 2. Figure S4. Correlation between the 53BP1 foci expression and tumor staging (see Additional file 1: Table S2). Peripheral lymphocytes were prepared from the blood samples derived from RC patients. Foci counting for 53BP1 were performed in non-irradiated (A), irradiated in vitro with 0.5 and 2 Gy samples 30 min and 24 h post-IR (B and C) or 72 h after 5 clinical radiation fractions (D). Filled squares represent the mean values (± SE) for the respective group. Figure S5. Comparison of the γ-H2AX foci expression in peripheral lymphocytes of RC patients differing in tumor regression grade (TRG, Additional file 1: Table S2). Foci counting for γ-H2AX were performed in non-irradiated (up triangles and circles) cells or after 5 clinical radiation fractions (down triangles and diamonds). Filled squares represent the mean values (± SE) for the respective group. (DOC 1915 kb

CIP2A is required for growth of HCT116 cells.

<p>(<b>A</b>) Immunoblot analysis of CIP2A and c-Myc protein expression in HCT116 cells infected lentiviral with shRNA targeting CIP2A or a ctr. shRNA. Numbers below lines indicate the c-myc protein expression relative to c-myc levels in control cells (n = 2). (<b>B</b>) Colony formation of HCT116 cells after 7 days. (<i>Top</i>), density of colonies stained with crystal violet; (<i>bottom</i>), representative of the indicated cultures (n = 3).</p

<i>CIP2A</i> mRNA expression is significantly correlated with advanced tumour stage.

<p>The panels show box and whisker plots documenting relative <i>CIP2A</i> mRNA levels in tumors stratified according to UICC stage (A) (UICC I vs. II p<.0001; II vs. III p = .0046; III vs. IV p = .0002), according to lymph node metastasis (B; N- vs. N+ p<.0001), according to distant metastasis (C; M0 vs. M1 p<.0001), and according to histological grading (D: G2 vs. G3 p = .0084).</p

CIP2A expression is regulated by MAPK signalling.

<p>Caco2, HCT116 and SW620 cells were treated with DMSO or the MEK inhibitor UO126 for 24(n = 3 for each cell line). (<b>A</b>) Immunoblot analysis of CIP2A and p-ERK protein expression in Caco2, HCT116 and SW620. (<b>B</b>) Real-time PCR analysis of <i>CIP2A</i> mRNA expression (*<0.05; **<0.005).</p

Depletion of CIP2A downregulates c-Myc protein expression in colon cancer cells.

<p>(<b>A</b>) Immunoblot analysis of CIP2A and c-Myc protein expression in Caco2, HCT116 and SW620 cells transfected with siRNA targeting CIP2A or control siRNA. Cells were harvested 72 h after transfection (n = 3 for each cell line). (<b>B</b>) Real-time PCR analysis of <i>CIP2A</i> and <i>c-Myc</i> mRNA expression in HCT116 cells transfected with siRNA targeting CIP2A or control siRNA (n = 3). (<b>C</b>) Depletion of CIP2A does not change activation status of AKT or its downstream targets. The panels show immunoblots of the indicated proteins and phosphorylated proteins (p) in Caco2, HCT116 and SW620 cells transfected with siRNA targeting CIP2A or control siRNA as before (n = 3 for each cell line).</p

CIP2A protein levels in colon cancer correlate with <i>CIP2A</i> mRNA expression.

<p>The panels show representative examples of immunofluorescence staining, showing CIP2A protein expression in cancer cells of patients with low (A+B) or high <i>CIP2A</i> (C+D) mRNA expression (A+C x100, B+D x200 magnification).</p