Survivin drives tumor-associated macrophage reprogramming: a novel mechanism with potential impact for obesity.

PURPOSE: Recent studies point to adipose-derived stem cells (ASCs) as a link between obesity and cancer. We aimed to determine whether survivin, which is highly secreted by ASCs from subjects with obesity, might drive a pro-tumoral phenotype in macrophages. METHODS: The effect of ASC conditioned medium on the macrophage phenotype was assessed by expression studies. Survivin intracellular localization and internalization were examined by subcellular fractionation and immunofluorescence, respectively. Loss- and gain-of-function studies were performed using adenoviral vectors, and gene expression patterns, migration and invasion capacities of cancer cells were examined. Heterotypic cultures of ASCs, macrophages and cancer cells were established to mimic the tumor microenvironment. Survivin-blocking experiments were used to determine the impact of survivin on both macrophages and cancer cells. Immunohistochemical analysis of survivin was performed in macrophages from ascitic fluids of cancer patients and healthy controls. RESULTS: We found that obese-derived ASCs induced a phenotypic switch in macrophages characterized by the expression of both pro- and anti-inflammatory markers. Macrophages were found to internalize extracellular survivin, generating hybrid macrophages with a tumor-associated phenotype that included secretion of survivin. Exogenous expression of survivin in macrophages generated a similar phenotype and enhanced the malignant characteristics of cancer cells by a mechanism dependent on survivin phosphorylation at threonine 34. Survivin secreted by both ASCs from subjects with obesity and tumor-associated macrophages synergistically boosted the malignancy of cancer cells. Importantly, survivin was mainly detected in ascites-associated macrophages from patients with a malignant diagnosis. CONCLUSION: Our data indicate that survivin may serve as a molecular link between obesity and cancer and as a novel marker for tumor-associated macrophages.

Wnt9a deficiency discloses a repressive role of Tcf7l2 on endocrine differentiation in the embryonic pancreas.

Transcriptional and signaling networks establish complex cross-regulatory interactions that drive cellular differentiation during development. Using microarrays we identified the gene encoding the ligand Wnt9a as a candidate target of Neurogenin3, a basic helix-loop-helix transcription factor that functions as a master regulator of pancreatic endocrine differentiation. Here we show that Wnt9a is expressed in the embryonic pancreas and that its deficiency enhances activation of the endocrine transcriptional program and increases the number of endocrine cells at birth. We identify the gene encoding the endocrine transcription factor Nkx2-2 as one of the most upregulated genes in Wnt9a-ablated pancreases and associate its activation to reduced expression of the Wnt effector Tcf7l2. Accordingly, in vitro studies confirm that Tcf7l2 represses activation of Nkx2-2 by Neurogenin3 and inhibits Nkx2-2 expression in differentiated ß-cells. Further, we report that Tcf7l2 protein levels decline upon initiation of endocrine differentiation in vivo, disclosing the downregulation of this factor in the developing endocrine compartment. These findings highlight the notion that modulation of signalling cues by lineage-promoting factors is pivotal for controlling differentiation programs.

Laparoscopy After Liver Transplant: Result of an Initial Series.

UNLABELLED: There are few reports about laparoscopic approach after liver transplantation (LT). The aim of this paper is to analyze in terms of feasibility and therapeutic effectiveness an initial experience with laparoscopy in patients who have had a liver transplantation. METHODS: Laparoscopic procedures were divided into "early" and "late" laparoscopy. RESULTS: A total of 10 laparoscopic procedures after LT were performed in 9 patients: 3 early and 7 late. The early laparoscopic procedures were performed in 3 patients who had a torpid evolution in their liver transplant: 2 were diagnostic only and 1 was also therapeutic. There were no intraoperative events or conversions. We recorded 2 complications. In 7 patients a late laparoscopy was performed: 4 in the upper abdomen and 3 in the lower abdomen. Of the 4 in the upper abdomen, 3 were liver procedures and 1 was in the transverse colon, on the other side; in addition, 2 laparoscopic inguinal hernia repairs and 1 laparoscopic appendectomy also were performed. None of these 7 cases was converted, and there were no intraoperative events. CONCLUSIONS: The early postoperative period, in critically ill patients diagnostic and eventually therapeutic laparoscopy was feasible and effective in our cases. Late laparoscopy in the upper abdomen is difficult due to adhesions and adds to the risk of poor graft function, but was feasible for both diagnostic and therapeutic purposes. Late laparoscopy in the lower abdomen with good graft function has the same safety and efficacy as those performed on nontransplanted patients.

Study of allelic losses on 3p, 6q, and 17p in human urothelial cancer.

Forty-eight transitional cell carcinomas of the bladder and three transitional cell carcinomas of the renal pelvis were examined for loss of heterozygosity (LOH) on chromosomes 3p, 6q, and 17p. The most frequent allelic loss was seen on 17p (18/36, 50%) followed by 6q (6/22, 27%), and 3p (5/22, 23%). In cases with LOH at more than one locus, the same DNA sample often varied in degree of signal reduction for missing alleles. This observation indicates that LOH studies can serve to detect intratumor heterogeneity. No correlation was found between allelic losses at these chromosome arms and tumor grade and stage. Allelic losses on 6q were associated with tumors having a solid growth pattern; in this kind of tumors, allelic losses on 3p were associated with invasion.

Analysis of 3p allelic losses in renal cell carcinomas: comparison with cytogenetic results.

We performed a study of loss of heterozygosity (LOH) at 3p by restriction fragment length polymorphism analysis in a series of 22 renal tumors. In 11 cases, molecular results could be compared with those of cytogenetic studies. The highest frequency of allelic losses at 3p was seen in clear cell non-papillary renal tumors, whereas none of the papillary renal cell carcinomas showed LOH at 3p. Allelic losses on 3p were found to be independent of tumor grade or stage or both. One case analyzed showed a discrepancy between cytogenetic and LOH studies. This tumor displayed rearrangements of chromosome 3 and no LOH at the c-RAF-1 (close to the Von Hippel Lindau gene) locus.

[Low incidence of mutations in codon 12 of the c-K-ras gene in bladder cancer]. / Baja incidencia de mutación del codon 12 del gen c-K-ras en el cáncer vesical.

During vesical carcinogenesis a variety of genetic alterations such as oncogene mutation or loss of suppressor genes have been detected. Codon 12 mutation of the c-K-ras gene has been seen with a high frequency in several human neoplasias but its participation in the development of vesical cancers has not been fully dilucidated. Using the DNA restriction fragments polymorphism (RFLP) technique enhanced by a polymerase chain reaction (PCR) a study has been made of codon 12 mutation at the c-K-ras gene in 55 patient with vesical cancer undergoing surgery between 1991 and 1992. The tumoral stage was superficial (Ta-Tl) in 24 cases, infiltrant (T2-T4) in 28 cases and unknown in 3 cases. Two patients (3.6%) showed codon 12 mutation at the c-K-ras gene. One case was a fast evolving infiltrant tumour (T2-T3) which caused death of the patient after 4 months while the other case was a surface tumour (G2Ta) which relapsed early, the pathological anatomy revealing a stage T2-T3 squamous carcinoma. Our results suggest that codon 12 mutation at the c-K-ras gene is not a meaningful genetic change in the genesis of vesical cancer. Its emergence, however, appears to be related to a more aggressive tumoural behaviour.

Restriction fragment length polymorphism of the L-myc gene is not a prognostic factor in bladder cancer patients.

The L-myc restriction fragment length polymorphism has been suggested to be of prognostic significance in some types of primary tumours. We examined the prognostic and susceptibility significance of the L-myc genotype in a group of 98 bladder cancer patients. The L-myc genotype did not correlate with any pathologic parameter and does not offer any clinical utility in patients with bladder cancer.

Clinical application of multiplex quantitative fluorescent polymerase chain reaction (QF-PCR) for the rapid prenatal detection of common chromosome aneuploidies.

The clinical application of quantitative fluorescent polymerase chain reaction (QF-PCR) for rapid prenatal detection of chromosome aneuploidies has been limited in most studies to the detection of autosomal trisomies. Recently it has been shown that a newly identified highly polymorphic marker, termed X22, which maps to the Xq/Yq pseudoautosomal region of the sex chromosomes, used together with the X-linked short tandem repeat (STR) HPRT, allows the accurate detection of gonosome aneuploidies. We have developed a rapid assay, which includes these STR markers together with a sequence of the amelogenin region of the sex chromosomes and selected highly polymorphic autosomal STR. Two more X chromosome markers, as yet not used in previous QF-PCR applications, were also included in the assay. The molecular test was then used in a clinical trial on 551 uncultured amniotic fluid samples, allowing the assessment of copy number for chromosomes X, Y and 21 in 100% of cases. In the course of this study, two fetuses with Turner's syndrome and one with Klinefelter's syndrome were identified along with 17 autosomal trisomies. The assay proved to be so efficient and reliable that in most aneuploidy cases, in which ultrasound findings were in agreement with the molecular result, therapeutical interventions were possible without waiting for the result of cytogenetic analysis.

Rapid prenatal diagnosis of common chromosome aneuploidies by QF-PCR. Assessment on 18,000 consecutive clinical samples.

The quantitative fluorescent PCR (QF-PCR) assay, introduced during the last few years, allows prenatal diagnoses of common chromosome aneuploidies in a few hours after sampling. We report the first assessment of QF-PCR performed on a large cohort of 18,000 consecutive clinical specimens analysed in two different Centres. All samples were analysed by QF-PCR using several selected STR markers together with amelogenin and, occasionally, SRY for fetal sexing. Results were compared with those obtained by conventional cytogenetic analysis. In 17,129 tests, normal fetuses were detected by QF-PCR. No false positives were observed. All 732 cases of trisomy 21, 18, 13, triploidies, double trisomies as well as all but one fetuses with X and Y aneuploidies were correctly diagnosed. Chromosome mosaicism could also be suspected in several samples. In some cases of in vitro culture failures, QF-PCR was the only evidence of fetal X, Y, 21, 18 and 13 chromosome complement. QF-PCR proved to be efficient and reliable in detecting major numerical chromosome disorders. The main advantages of the molecular assay are its very low cost, speed and automation enabling a single operator to perform up to 40 assays per day. QF-PCR relieves anxiety of most parents within 24 h from sampling and accelerates therapeutic interventions in the case of an abnormal result. In countries where large scale conventional cytogenetics is hampered by its high cost and lack of technical expertise, QF-PCR may be used as the only prenatal diagnostic test.