Autocrine CSF-1R signaling drives mesothelioma chemoresistance via AKT activation.

Clinical management of malignant pleural mesothelioma (MPM) is very challenging because of the uncommon resistance of this tumor to chemotherapy. We report here increased expression of macrophage colony-stimulating-factor-1-receptor (M-CSF/CSF-1R) mRNA in mesothelioma versus normal tissue specimens and demonstrate that CSF-1R expression identifies chemoresistant cells of mesothelial nature in both primary cultures and mesothelioma cell lines. By using RNAi or ligand trapping, we demonstrate that the chemoresistance properties of those cells depend on autocrine CSF-1R signaling. At the single-cell level, the isolated CSF-1R(pos) cells exhibit a complex repertoire of pluripotency, epithelial-mesenchymal transition and detoxifying factors, which define a clonogenic, chemoresistant, precursor-like cell sub-population. The simple activation of CSF-1R in untransformed mesothelial cells is sufficient to confer clonogenicity and resistance to pemetrexed, hallmarks of mesothelioma. In addition, this induced a gene expression profile highly mimicking that observed in the MPM cells endogenously expressing the receptor and the ligands, suggesting that CSF-1R expression is mainly responsible for the phenotype of the identified cell sub-populations. The survival of CSF1R(pos) cells requires active AKT (v-akt murine thymoma viral oncogene homolog 1) signaling, which contributed to increased levels of nuclear, transcriptionally competent ß-catenin. Inhibition of AKT reduced the transcriptional activity of ß-catenin-dependent reporters and sensitized the cells to senescence-induced clonogenic death after pemetrexed treatment. This work expands what is known on the non-macrophage functions of CSF-1R and its role in solid tumors, and suggests that CSF-1R signaling may have a critical pathogenic role in a prototypical, inflammation-related cancer such as MPM and therefore may represent a promising target for therapeutic intervention.

Protumorigenic effects of mir-145 loss in malignant pleural mesothelioma.

We identified a discrete number of microRNAs differentially expressed in benign or malignant mesothelial tissues. We focused on mir-145 whose levels were significantly downregulated in malignant mesothelial tissues and malignant pleural mesothelioma (MPM) cell lines as compared to benign tissues (pleura, peritoneum or cysts). We show that promoter hyper-methylation caused very low levels in MPM cell lines and specimens. Treatment of MPM cell lines with mir-145 agonists negatively modulated some protumorigenic properties of MPM cells, such as clonogenicity, cell migration and resistance to pemetrexed treatment. The main effector mechanism of the clonogenic death induced by mir-145 was that of accelerated senescence. We found that mir-145 targeted OCT4 via specific binding to its 3'-UTR. Increased intracellular levels of mir-145 decreased the levels of OCT4 and its target gene ZEB1, thereby counteracting the increase of OCT4 induced by pemetrexed treatment which is known to favor the development of chemoresistant cells. In line with this, reintroduction of OCT4 into mimic-145 treated cells counteracted the effects on clonogenicity and replicative senescence. This further supports the relevance of the mir-145-OCT4 interaction for the survival of MPM cells. The potential use of mir-145 expression levels to classify benign vs malignant mesothelial tissues and the differences between pemetrexed-induced senescence and that induced by the re-expression of mir-145 are discussed.

SASP mediates chemoresistance and tumor-initiating-activity of mesothelioma cells.

Here we show that pemetrexed-treated mesothelioma cells undergo accelerated senescence. This is characterized by the secretion of proinflammatory and mitogenic cytokines, reminiscent of an SASP (senescence-associated secretory phenotype). Conditioned media from senescent MPM (malignant pleural mesothelioma) cells trigger the emergence of EMT (epithelial-to-mesenchymal)-like, clonogenic and chemoresistant cell subpopulations, expressing high levels of ALDH (aldehyde dehydrogenase) activity (ALDH(bright) cells). We show by fluorescence-activated cell sorting of purified ALDH(bright) and ALDH(low) cells, that both cell-autonomous and cell-non-autonomous mechanisms converge to maintain the SASP-induced, EMT-like cell subpopulations. Chemoresistant ALDH(bright) cells exist within primary MPM specimens and enrichment for ALDH(bright) cells correlates with an earlier tumor onset into NOD/SCID mice. We show that RAS(v12) expression induces SASP-like changes in untransformed human mesothelial cells, and that p53 ablation increases the effect of RAS(v12) expression. We identify STAT3 activation as a crucial event downstream to SASP signaling. In fact, small hairpin RNA-mediated ablation of STAT3 deeply attenuates the induction of EMT genes and the increase of ALDH(bright) cells induced by SASP-cytokines. This strongly affects the chemoresistance of MPM cells in vitro and leads to anticancer effects in vivo.

Cytotoxic proteins combined with prodigiosin obtained from Serratia marcescens have both broad and selective cytotoxic activity on tumor cells.

Cytotoxic proteins and prodigiosin obtained from Serratia marcescens strains are known to induce tumor cell death, nevertheless its combination has not been studied. In this paper we evaluate the combined effects of these molecules in a panel of tumor cell lines. The results showed a marked inhibitory effect on the growth of tumor cell lines derived from tumors (i.e., melanoma) which are highly resistant to conventional anticancer drugs, while normal cells were less sensitive than tumor cells. TUNEL (TdT-mediated dUTP nick end labeling) and electrophoresis of HEp-2 cell DNA treated with MG2327 preparation [containing the P50 protein belonging to the serralysins and prodigiosin, from S. marcescens CMIB4202] showed a pattern of DNA fragments typically associated with apoptosis. Interestingly, prodigiosin enhanced by 1.6-fold the cytotoxic effect of P50 when acting in combination on HEp-2 cells. The broad cytotoxic activity of the combination on tumor cells as well as its selectivity open new frontiers in cancer therapy.

Early childhood surveillance of developmental disorders by a birth defects surveillance system: methods, prevalence comparisons, and mortality patterns.

The prevalence of developmental disabilities in early childhood is not well documented. An established birth defects registry extended surveillance to identify cases of developmental disorders in early childhood by adding all known sources of diagnosis and service to case-finding methods. Residents of a northwest Arkansas region born during 1985 to 1987 and diagnosed with either a birth defect or a developmental disorder by the 4th birthday comprised the studied cohort. Case records were linked with death certificates to examine the influence of mortality on prevalence ratios. Prevalence ratios estimated were 64.5/1000 resident live births (60.9/1000 among survivors to age 4 years) for either birth defect or developmental disorder, 33.4/1000 for developmental disorder, 37.0/1000 for birth defect, and 9.5/1000 for both developmental disorder and birth defect. Prevalence ratios of specific developmental disorders and the role of mortality in decreasing population prevalence are reported. The most common diagnostic categories in this age group were developmental delay, seizures, and failure to thrive. Overlap of birth defect categories with a diagnosed developmental disability was examined; 68.8% of children diagnosed with neural tube defects and 45.5% of those with chromosomal abnormalities who survived to age 4 years had clinically diagnosed developmental disorders. An anticipated high degree of overlap (77%) for other central nervous system defects was found. For other birth defect categories, developmental disorder diagnosis was present in 20 to 30% of the study group. Death before age 4 years occurred most often when the diagnosis was newborn seizures (17.1%) or "conditions of the brain" (13.6%); the mortality rate was 6 to 8% for epilepsy or seizure disorders, mental retardation, and vision loss. The large number of developmental diagnoses among this cohort indicates that surveillance of these disorders in early childhood, even with tentative diagnoses, is feasible. Data obtained indicate that many birth defects are associated with developmental disorders; potentially, this association can contribute to earlier identification of developmental disorders in childhood.

Deaths associated with renal agenesis: a population-based study of birth prevalence, case ascertainment, and etiologic heterogeneity.

We report on deaths associated with renal agenesis among 211,704 consecutive births. Sources included birth and death certificates and an active birth defects surveillance system. Medical review and classification of cases were performed for 1985-1990 events. Sixty-one cases of renal agenesis were identified, and review of records was possible for 59 of the 61 cases. Of these 59 cases, 36 (61%) were confirmed, 5 (8%) were questionable, and 18 (31%) were incorrectly coded. The prevalence of confirmed cases is thus estimated at 17/100,000 births (14.2/100,000 births, excluding elective terminations and fetal deaths). Records incorrectly coded were most often those with multicystic dysplasia. Approximately one-third of cases was found by the birth defects surveillance system alone, confirming the utility of this data source for prevalence estimates. Isolated renal agenesis accounted for 44% of confirmed cases; other diagnoses included VATER association (19%), unrecognized multiple malformation syndromes (17%), exstrophy of the cloaca sequence (14%), and chromosome disorders (6%). Based on these data, prevalence rates for ICD code 753.0 and death include overascertainment of cases from erroneous coding of multicystic dysplasia and underascertainment of cases with unilateral renal agenesis associated with other malformations. Population-based ascertainment of cases by active surveillance methods and rigorous diagnostic coding standards are required to improve the accuracy of these rates. Targeted investigations of distinct subclassifications will be necessary to identify specific etiologic factors.

Evaluation of renal transplant rejection by duplex Doppler examination: value of the resistive index.

The increasing use and availability of renal transplantation has resulted in a demand for noninvasive methods to study possible complications. One of the most serious adverse reactions is acute rejection, a possibly reversible cause of transplant failure if treated promptly. Differentiation from other causes of acute renal failure frequently is difficult, and the lack of specificity in many imaging studies has been troublesome. Eighty-one patients with renal transplants, including 41 with acute rejection, were examined. Duplex Doppler examination of the intrarenal arteries and a simplified formula, the resistive index ([peak systolic frequency shift--lowest diastolic frequency shift]/[peak systolic frequency shift]), were used to diagnose rejection. With a resistive index greater than 0.90, a 100% positive predictive value was obtained for the diagnosis of acute rejection. A value less than 0.70 was unlikely to be rejection (negative predictive value, 94%). This approach uses a simple analysis of the waveform. Use of a duplex Doppler examination and the formula described here appears to be an accurate method for the detection of acute rejection and for the differentiation of acute rejection from the various other causes of acute renal failure.

Double contrast knee arthrography in children.

The indications for knee arthrography in children consist of: suspected internal derangement; osteochondritis dissecans; Blount's disease; popliteal cysts; loose bodies; synovial tumors; and miscellaneous abnormalities including unexplained knee pain.