Single-Cell Transcriptomics of Human and Mouse Lung Cancers Reveals Conserved Myeloid Populations across Individuals and Species.

Tumor-infiltrating myeloid cells (TIMs) comprise monocytes, macrophages, dendritic cells, and neutrophils, and have emerged as key regulators of cancer growth. These cells can diversify into a spectrum of states, which might promote or limit tumor outgrowth but remain poorly understood. Here, we used single-cell RNA sequencing (scRNA-seq) to map TIMs in non-small-cell lung cancer patients. We uncovered 25 TIM states, most of which were reproducibly found across patients. To facilitate translational research of these populations, we also profiled TIMs in mice. In comparing TIMs across species, we identified a near-complete congruence of population structures among dendritic cells and monocytes; conserved neutrophil subsets; and species differences among macrophages. By contrast, myeloid cell population structures in patients' blood showed limited overlap with those of TIMs. This study determines the lung TIM landscape and sets the stage for future investigations into the potential of TIMs as immunotherapy targets.

mRNA circularization by METTL3-eIF3h enhances translation and promotes oncogenesis.

N6-methyladenosine (m6A) modification of mRNA is emerging as an important regulator of gene expression that affects different developmental and biological processes, and altered m6A homeostasis is linked to cancer1-5. m6A modification is catalysed by METTL3 and enriched in the 3' untranslated region of a large subset of mRNAs at sites close to the stop codon5. METTL3 can promote translation but the mechanism and relevance of this process remain unknown1. Here we show that METTL3 enhances translation only when tethered to reporter mRNA at sites close to the stop codon, supporting a mechanism of mRNA looping for ribosome recycling and translational control. Electron microscopy reveals the topology of individual polyribosomes with single METTL3 foci in close proximity to 5' cap-binding proteins. We identify a direct physical and functional interaction between METTL3 and the eukaryotic translation initiation factor 3 subunit h (eIF3h). METTL3 promotes translation of a large subset of oncogenic mRNAs-including bromodomain-containing protein 4-that is also m6A-modified in human primary lung tumours. The METTL3-eIF3h interaction is required for enhanced translation, formation of densely packed polyribosomes and oncogenic transformation. METTL3 depletion inhibits tumorigenicity and sensitizes lung cancer cells to BRD4 inhibition. These findings uncover a mechanism of translation control that is based on mRNA looping and identify METTL3-eIF3h as a potential therapeutic target for patients with cancer.

Large-scale analysis of BAP1 expression reveals novel associations with clinical and molecular features of malignant pleural mesothelioma.

BRCA1-associated protein-1 (BAP1) expression is commonly lost in several tumors including malignant pleural mesothelioma (MPM). Presence or absence of immunohistochemical BAP1 nuclear staining in tumor cells is currently used for differential diagnosis of MPM. In this study, a large cohort of 596 MPM tumors with available clinical data was analyzed to examine associations of BAP1 staining pattern with clinical and molecular features that may reflect the impact of BAP1 mutation on MPM biology. Cases were classified according to the BAP1 staining pattern of tumor cells. Exome and RNA-sequencing data were available for subsets of cases. Levels of mRNA encoding claudin 15 (CLDN15) and vimentin (VIM) were determined using RT-qPCR on 483 cases to estimate the relative proportions of epithelial-like and mesenchymal-like components in each tumor. Four BAP1 staining patterns were observed: single-pattern nuclear staining (36%), single-pattern cytoplasmic staining (25%), single-pattern absent staining (12%), and combinations of these staining patterns (27%). This study confirmed prior reports that nuclear BAP1 is more frequently associated with wild-type BAP1 and sarcomatoid histology. However, no associations between BAP1 staining pattern(s) and mutations in specific protein domains and/or mutation type were observed. BAP1 staining patterns were significantly associated (p < 0.001) with BAP1 gene expression, MPM histologic subtypes, molecular clusters, and markers of epithelial-to-mesenchymal transition. Frequent observation of combinations of BAP1 staining patterns in MPM tumors indicated intra-tumoral heterogeneity of BAP1 status. Cytoplasmic BAP1 staining was identified as a putative indicator of favorable prognosis in non-epithelioid MPM. In conclusion, novel significant associations among different BAP1 staining patterns and subgroups of MPM tumors were observed, suggesting that the role of BAP1 in tumor progression may be more complex than its presumed tumor suppressor function. Cytoplasmic staining was identified as a putative indicator of favorable prognosis in non-epithelioid MPM, potentially addressing a critical need in clinical decision-making in this disease. © 2020 The Authors. The Journal of Pathology published by John Wiley & Sons, Ltd. on behalf of The Pathological Society of Great Britain and Ireland.

A biparatopic agonistic antibody that mimics fibroblast growth factor 21 ligand activity.

Bispecific antibodies have become important formats for therapeutic discovery. They allow for potential synergy by simultaneously engaging two separate targets and enable new functions that are not possible to achieve by using a combination of two monospecific antibodies. Antagonistic antibodies dominate drug discovery today, but only a limited number of agonistic antibodies (i.e. those that activate receptor signaling) have been described. For receptors formed by two components, engaging both of these components simultaneously may be required for agonistic signaling. As such, bispecific antibodies may be particularly useful in activating multicomponent receptor complexes. Here, we describe a biparatopic (i.e. targeting two different epitopes on the same target) format that can activate the endocrine fibroblast growth factor (FGF) 21 receptor (FGFR) complex containing ß-Klotho and FGFR1c. This format was constructed by grafting two different antigen-specific VH domains onto the VH and VL positions of an IgG, yielding a tetravalent binder with two potential geometries, a close and a distant, between the two paratopes. Our results revealed that the biparatopic molecule provides activities that are not observed with each paratope alone. Our approach could help address the challenges with heterogeneity inherent in other bispecific formats and could provide the means to adjust intramolecular distances of the antibody domains to drive optimal activity in a bispecific format. In conclusion, this format is versatile, is easy to construct and produce, and opens a new avenue for agonistic antibody discovery and development.

Calcimimetics maintain bone turnover in uremic rats despite the concomitant decrease in parathyroid hormone concentration.

Calcimimetics decrease parathyroid hormone (PTH) secretion in patients with secondary hyperparathyroidism. The decrease in PTH should cause a reduction in bone turnover; however, the direct effect of calcimimetics on bone cells, which express the calcium-sensing receptor (CaSR), has not been defined. In this study, we evaluated the direct bone effects of CaSR activation by a calcimimetic (AMG 641) in vitro and in vivo. To create a PTH "clamp," total parathyroidectomy was performed in rats with and without uremia induced by 5/6 nephrectomy, followed by a continuous subcutaneous infusion of PTH. Animals were then treated with either the calcimimetic or vehicle. Calcimimetic administration increased osteoblast number and osteoid volume in normal rats under a PTH clamp. In uremic rats, the elevated PTH concentration led to reduced bone volume and increased bone turnover, and calcimimetic administration decreased plasma PTH. In uremic rats exposed to PTH at 6-fold the usual replacement dose, calcimimetic administration increased osteoblast number, osteoid surface, and bone formation. A 9-fold higher dose of PTH caused an increase in bone turnover that was not altered by the administration of calcimimetic. In an osteosarcoma cell line, the calcimimetic induced Erk1/2 phosphorylation and the expression of osteoblast genes. The addition of a calcilytic resulted in the opposite effect. Moreover, the calcimimetic promoted the osteogenic differentiation and mineralization of human bone marrow mesenchymal stem cells in vitro. Thus, calcimimetic administration has a direct anabolic effect on bone that counteracts the decrease in PTH levels.

Diagnostic value of biopsy sampling in predicting histology in patients with diffuse malignant pleural mesothelioma.

BACKGROUND: The classification of diffuse malignant mesothelioma into epithelioid, biphasic, and sarcomatoid types is based on histologic patterns. The diagnosis is made on biopsies, and because of intratumoral heterogeneity, they may not be representative of the entire tumor. The number and volume of biopsies needed to reach diagnostic accuracy in diffuse malignant mesothelioma and their prognostic value remain unclear. METHODS: This study examined 759 consecutive patients with pleural diffuse malignant mesothelioma treated by pleurectomy/decortication or extrapleural pneumonectomy for the presence of epithelioid and/or sarcomatoid histology and classified both the presurgery biopsies (core-needle or thoracoscopic) and surgical resection specimens. The number and volume of biopsies were correlated with pre- and postsurgery histologies and overall survival. RESULTS: Diffuse malignant mesothelioma was classified as epithelioid (76%), biphasic (18%), sarcomatoid (5%), or indeterminate (1%) in biopsies and as epithelioid (64%), biphasic (32%), and sarcomatoid (4%) in surgical resection specimens (overall concordance, 80.6%). The positive likelihood ratios were 2.4, 13.6, and 90.1 for biopsies with epithelioid, biphasic, and sarcomatoid histologies, respectively. Concordant histologies between biopsies and resections were associated with a higher number of biopsies (median tissue blocks for concordant histologies vs discordant histologies, 3 vs 2; P < .002) but were less associated with a higher volume (median, 1.2 vs 1.1 cm3 ; P = .06). In a multivariate analysis, overall survival was independently predicted by histology in the resection specimen (P < .0001) but not in the biopsy (P = .09). CONCLUSIONS: In contrast to epithelioid histology, sarcomatoid histology in biopsies is highly accurate. Despite intratumoral heterogeneity, the accuracy of histologic classification increases with the number of tissue blocks examined, emphasizing the diagnostic value of extensive sampling by presurgery biopsies.

Autophagy facilitates the release of immunogenic signals following chemotherapy in 3D models of mesothelioma.

We have previously shown that nearly half of mesothelioma patients have tumors with low autophagy and that these patients have a significantly worse outcome than those with high autophagy. We hypothesized that autophagy may be beneficial by facilitating immunogenic cell death (ICD) of tumor cells following chemotherapy. An important hallmark of ICD is that death of tumor cells is preceded or accompanied by the release of damage-associated molecular pattern molecules (DAMPs), which then can stimulate an antitumor immune response. Therefore, we measured how autophagy affected the release of three major DAMPs: high mobility group box 1 (HMGB1), ATP, and calreticulin following chemotherapy. We found that autophagy in three-dimensional (3D) models with low autophagy at baseline could be upregulated with the cell-permeant Tat-BECN1 peptide and confirmed that autophagy in 3D models with high autophagy at baseline could be inhibited with MRT 68921 or ATG7 RNAi, as we have previously shown. In in vitro 3D spheroids, we found that, when autophagy was high or upregulated, DAMPs were released following chemotherapy; however, when autophagy was low or inhibited, DAMPs release was significantly impaired. Similarly, in ex vivo tumors, when autophagy was high or upregulated, HMGB1 was released following chemotherapy but, when autophagy was low, HMGB1 release was not seen. We conclude that autophagy can be upregulated in at least some tumors with low autophagy and that upregulation of autophagy can restore the release of DAMPs following chemotherapy. Autophagy may be necessary for ICD in this tumor.

In Vitro and in Vivo Analyses Reveal Profound Effects of Fibroblast Growth Factor 16 as a Metabolic Regulator.

The discovery of brown adipose tissue (BAT) as a key regulator of energy expenditure has sparked interest in identifying novel soluble factors capable of activating inducible BAT (iBAT) to combat obesity. Using a high content cell-based screen, we identified fibroblast growth factor 16 (FGF16) as a potent inducer of several physical and transcriptional characteristics analogous to those of both "classical" BAT and iBAT. Overexpression of Fgf16 in vivo recapitulated several of our in vitro findings, specifically the significant induction of the Ucp1 gene and UCP1 protein expression in inguinal white adipose tissue (iWAT), a common site for emergent active iBAT. Despite significant UCP1 up-regulation in iWAT and dramatic weight loss, the metabolic improvements observed due to Fgf16 overexpression in vivo were not the result of increased energy expenditure, as measured by indirect calorimetric assessment. Instead, a pattern of reduced food and water intake, combined with feces replete with lipid and bile acid, indicated a phenotype more akin to that of starvation and intestinal malabsorption. Gene expression analysis of the liver and ileum indicated alterations in several steps of bile acid metabolism, including hepatic synthesis and reabsorption. Histological analysis of intestinal tissue revealed profound abnormalities in support of this conclusion. The in vivo data, together with FGF receptor binding analysis, indicate that the in vivo outcome observed is the likely result of both direct and indirect mechanisms and probably involves multiple receptors. These results highlight the complexity of FGF signaling in the regulation of various metabolic processes.

FGF23 impairs peripheral microvascular function in renal failure.

Cardiovascular diseases account for ~50% of mortality in patients with chronic kidney disease (CKD). Fibroblast growth factor 23 (FGF23) is independently associated with endothelial dysfunction and cardiovascular mortality. We hypothesized that CKD impairs microvascular endothelial function and that this can be attributed to FGF23. Mice were subjected to partial nephrectomy (5/6Nx) or sham surgery. To evaluate the functional role of FGF23, non-CKD mice received FGF23 injections and CKD mice received FGF23-blocking antibodies after 5/6Nx surgery. To examine microvascular function, myocardial perfusion in vivo and vascular function of gracilis resistance arteries ex vivo were assessed in mice. 5/6Nx surgery blunted ex vivo vasodilator responses to acetylcholine, whereas responses to sodium nitroprusside or endothelin were normal. In vivo FGF23 injections in non-CKD mice mimicked this endothelial defect, and FGF23 antibodies in 5/6Nx mice prevented endothelial dysfunction. Stimulation of microvascular endothelial cells with FGF23 in vitro did not induce ERK phosphorylation. Increased plasma asymmetric dimethylarginine concentrations were increased by FGF23 and strongly correlated with endothelial dysfunction. Increased FGF23 concentration did not mimic impaired endothelial function in the myocardium of 5/6Nx mice. In conclusion, impaired peripheral endothelium-dependent vasodilatation in 5/6Nx mice is mediated by FGF23 and can be prevented by blocking FGF23. These data corroborate FGF23 as an important target to combat cardiovascular disease in CKD. NEW & NOTEWORTHY In the present study, we provide the first evidence that fibroblast growth factor 23 (FGF23) is a cause of peripheral endothelial dysfunction in a model of early chronic kidney disease (CKD) and that endothelial dysfunction in CKD can be prevented by blockade of FGF23. This pathological effect on endothelial cells was induced by long-term exposure of physiological levels of FGF23. Mechanistically, increased plasma asymmetric dimethylarginine concentrations were strongly associated with this endothelial dysfunction in CKD and were increased by FGF23.

Inhibition of autophagy initiation potentiates chemosensitivity in mesothelioma.

The benefits of inhibiting autophagy in cancer are still controversial, with differences in outcome based on the type of tumor, the context and the particular stage of inhibition. Here, we investigated the impact of inhibiting autophagy at different stages on chemosensitivity using 3-dimensional (3D) models of mesothelioma, including ex vivo human tumor fragment spheroids. As shown by LC3B accumulation, we successfully inhibited autophagy using either an early stage ULK1/2 inhibitor (MRT 68921) or a late stage inhibitor (hydroxychloroquine). We found that inhibition of autophagy at the early stage, but not at late stage, potentiated chemosensitivity. This effect was seen only in those spheroids with high autophagy and active initiation at steady state. Inhibition of autophagy alone, at either early or late stage, did not cause cell death, showing that the inhibitors were non-toxic and that mesothelioma did not depend on autophagy at baseline, at least over 24 h. Using ATG13 puncta analysis, we found that autophagy initiation identified tumors that are more chemosensitive at baseline and after autophagy inhibition. Our results highlight a potential role of autophagy initiation in supporting mesothelioma cells during chemotherapy. Our work also highlights the importance of testing the inhibition of different stages in order to uncover the role of autophagy and the potential of its modulation in the treatment of cancer.

Differential regulation of renal Klotho and FGFR1 in normal and uremic rats.

In renal failure, hyperphosphatemia occurs despite a marked elevation in serum fibroblast growth factor (FGF)-23. Abnormal regulation of the FGFR1-Klotho receptor complex may cause a resistance to the phosphaturic action of FGF23. The purpose of the present study was to investigate the regulation of renal Klotho and FGF receptor (FEFR)-1 in healthy and uremic rats induced by 5/6 nephrectomy. In normal rats, the infusion of rat recombinant FGF23 enhanced phosphaturia and increased renal FGFR1 expression; however, Klotho expression was reduced. Uremic rats on a high-phosphate (HP) diet presented hyperphosphatemia with marked elevation of FGF23 and an increased fractional excretion of phosphate (P) that was associated with a marked reduction of Klotho expression and an increase in FGFR1. After neutralization of FGF23 by anti-FGF23 administration, phosphaturia was still abundant, Klotho expression remained low, and the FGFR1 level was reduced. These results suggest that the expression of renal Klotho is modulated by phosphaturia, whereas the FGFR1 expression is regulated by FGF23. Calcitriol (CTR) administration prevented a decrease in renal Klotho expression. In HEK293 cells HP produced nuclear translocation of ß-catenin, together with a reduction in Klotho. Wnt/ß-catenin inhibition with Dkk-1 prevented the P-induced down-regulation of Klotho. The addition of CTR to HP medium was able to recover Klotho expression. In summary, high FGF23 levels increase FGFR1, whereas phosphaturia decreases Klotho expression through the activation of Wnt/ß-catenin pathway.-Muñoz-Castañeda, J. R., Herencia, C., Pendón-Ruiz de Mier, M. V., Rodriguez-Ortiz, M. E., Diaz-Tocados, J. M., Vergara, N., Martínez-Moreno, J. M., Salmerón, M. D., Richards, W. G., Felsenfeld, A., Kuro-O, M., Almadén, Y., Rodríguez, M. Differential regulation of renal Klotho and FGFR1 in normal and uremic rats.

Etelcalcetide, A Novel Calcimimetic, Prevents Vascular Calcification in A Rat Model of Renal Insufficiency with Secondary Hyperparathyroidism.

Etelcalcetide, a novel peptide agonist of the calcium-sensing receptor, prevents vascular calcification in a rat model of renal insufficiency with secondary hyperparathyroidism. Vascular calcification occurs frequently in patients with chronic kidney disease (CKD) and is a consequence of impaired mineral homeostasis and secondary hyperparathyroidism (SHPT). Etelcalcetide substantially lowers parathyroid hormone (PTH) and fibroblast growth factor-23 (FGF23) levels in SHPT patients on hemodialysis. This study compared the effects of etelcalcetide and paricalcitol on vascular calcification in rats with adenine-induced CKD and SHPT. Uremia and SHPT were induced in male Wistar rats fed a diet supplemented with 0.75% adenine for 4 weeks. Rats were injected with vehicle, etelcalcetide, or paricalcitol for 4 weeks from the beginning of adenine diet. Rats fed an adenine-free diet were included as nonuremic controls. Similar reductions in plasma PTH and parathyroid chief cell proliferation were observed in both etelcalcetide- and paricalcitol-treated rats. Serum calcium and phosphorus were significantly lower in etelcalcetide-treated uremic rats and was unchanged in paricalcitol-treated rats. Both serum FGF23 and aortic calcium content were significantly lower in etelcalcetide-treated uremic rats compared with either vehicle- or paricalcitol-treated uremic rats. The degree of aortic calcium content for etelcalcetide-treated rats was similar to that in nonuremic controls and corroborated findings of lack of histologic aortic mineralization in those groups. In conclusion, etelcalcetide and paricalcitol similarly attenuated progression of SHPT in an adenine rat model of CKD. However, etelcalcetide differentially prevented vascular calcification, at least in part, due to reductions in serum FGF23, calcium, and phosphorus levels.

Transcriptome analysis reveals manifold mechanisms of cyst development in ADPKD.

BACKGROUND: Autosomal dominant polycystic kidney disease (ADPKD) causes progressive loss of renal function in adults as a consequence of the accumulation of cysts. ADPKD is the most common genetic cause of end-stage renal disease. Mutations in polycystin-1 occur in 87% of cases of ADPKD and mutations in polycystin-2 are found in 12% of ADPKD patients. The complexity of ADPKD has hampered efforts to identify the mechanisms underlying its pathogenesis. No current FDA (Federal Drug Administration)-approved therapies ameliorate ADPKD progression. RESULTS: We used the de Almeida laboratory's sensitive new transcriptogram method for whole-genome gene expression data analysis to analyze microarray data from cell lines developed from cell isolates of normal kidney and of both non-cystic nephrons and cysts from the kidney of a patient with ADPKD. We compared results obtained using standard Ingenuity Volcano plot analysis, Gene Set Enrichment Analysis (GSEA) and transcriptogram analysis. Transcriptogram analysis confirmed the findings of Ingenuity, GSEA, and published analysis of ADPKD kidney data and also identified multiple new expression changes in KEGG (Kyoto Encyclopedia of Genes and Genomes) pathways related to cell growth, cell death, genetic information processing, nucleotide metabolism, signal transduction, immune response, response to stimulus, cellular processes, ion homeostasis and transport and cofactors, vitamins, amino acids, energy, carbohydrates, drugs, lipids, and glycans. Transcriptogram analysis also provides significance metrics which allow us to prioritize further study of these pathways. CONCLUSIONS: Transcriptogram analysis identifies novel pathways altered in ADPKD, providing new avenues to identify both ADPKD's mechanisms of pathogenesis and pharmaceutical targets to ameliorate the progression of the disease.

Inhibition of secreted frizzled-related protein 5 improves glucose metabolism.

Elucidating the role of secreted frizzled-related protein 5 (SFRP5) in metabolism and obesity has been complicated by contradictory findings when knockout mice were used to determine metabolic phenotypes. By overexpressing SFRP5 in obese, prediabetic mice we consistently observed elevated hyperglycemia and glucose intolerance, supporting SFRP5 as a negative regulator of glucose metabolism. Accordingly, Sfrp5 mRNA expression analysis of both epididymal and subcutaneous adipose depots of mice indicated a correlation with obesity. Thus, we generated a monoclonal antibody (mAb) against SFRP5 to ascertain the effect of SFRP5 inhibition in vivo. Congruent with SFRP5 overexpression worsening blood glucose levels and glucose intolerance, anti-SFRP5 mAb therapy improved these phenotypes in vivo. The results from both the overexpression and mAb inhibition studies suggest a role for SFRP5 in glucose metabolism and pancreatic ß-cell function and thus establish the use of an anti-SFRP5 mAb as a potential approach to treat type 2 diabetes.

Parathyroid hormone activates the orphan nuclear receptor Nurr1 to induce FGF23 transcription.

Parathyroid hormone (PTH) increases FGF23 mRNA and protein levels in vivo and in vitro. Here we tested whether the increased FGF23 expression by PTH is mediated by the orphan nuclear receptor Nurr1. PTH increased Nurr1 mRNA levels prior to elevation of FGF23 mRNA levels in UMR-106 rat osteoblast-like cells. Activation of PKA increased both FGF23 and Nurr1 mRNA levels. Modification of Nurr1 expression showed that Nurr1 is essential for the PTH-mediated increase in FGF23 and luciferase reporter gene experiments identified a functional promoter region containing several potential Nurr1 binding sites. Chromatin immunoprecipitation assays confirmed the binding of Nurr1 to these regions in the FGF23 promoter. In vivo, Nurr1 mRNA and protein levels were increased in calvaria from rats with experimental CKD together with high PTH and FGF23 expression. Calcimimetics decrease PTH and FGF23 levels in CKD patients. Importantly, in rats with experimental CKD, the calcimimetic R568 decreased PTH expression, calvaria Nurr1 mRNA and protein levels, and FGF23 mRNA. Immunohistochemistry for Nurr1 showed an increase in the number of Nurr1 expressing osteocytes in the femurs of rats with CKD and this was decreased by R568. Thus, the effect of PTH to increase FGF23 transcription is mediated by Nurr1 in vitro and in vivo. In CKD, calcimimetics decrease PTH, which in turn decreases Nurr1 and consequently FGF23.

Extrapleural pneumonectomy in the treatment of epithelioid malignant pleural mesothelioma: novel prognostic implications of combined N1 and N2 nodal involvement based on experience in 529 patients.

OBJECTIVE: We review our 24-year experience with extrapleural pneumonectomy (EPP) in the treatment of epithelioid malignant pleural mesothelioma (MPM). BACKGROUND: Recent publications, particularly the MARS (Mesothelioma and Radical Surgery) feasibility study by Treasure et al, have questioned the safety and efficacy of EPP for MPM. METHODS: An institutional review board-approved, prospective, single-center database was retrospectively reviewed. Descriptive statistics and Kaplan-Meier analysis of overall survival are reported. RESULTS: From 1988 to 2011, a total of 529 patients with epithelioid MPM underwent complete resection by EPP as part of a multimodality strategy. Among these, 131 (25%) were women, and the median age was 59 (range, 17-79) years. Median postoperative hospital stay was 10 (range, 1-101) days. Twenty-six patients (5%) experienced 30-day or in-hospital mortality. Median overall survival was 18 months, with 1-, 3-, 5-, and 10-year survival rates of 67%, 28%, 14%, and 4%, respectively. Outcome by pathologic lymph node status (N, median overall survival) was N0: 224, 26 months; N1: 118, 17 months; N2: 181, 13 months; N3: 5, 7 months; Nx: 1, not evaluable. CONCLUSIONS: EPP has evolved as an effective method for macroscopic complete resection. This study confirms that lymph node status is significantly correlated with overall survival in patients with epithelioid MPM undergoing EPP and suggests that those with simultaneous involvement of N1 and N2 stations are at increased risk. This observation underscores the need for thorough staging of both N1 and N2 stations and has implications for revision of MPM staging criteria.

Dickkopf-1 is a master regulator of joint remodeling.

Degenerative and inflammatory joint diseases lead to a destruction of the joint architecture. Whereas degenerative osteoarthritis results in the formation of new bone, rheumatoid arthritis leads to bone resorption. The molecular basis of these different patterns of joint disease is unknown. By inhibiting Dickkopf-1 (DKK-1), a regulatory molecule of the Wnt pathway, we were able to reverse the bone-destructive pattern of a mouse model of rheumatoid arthritis to the bone-forming pattern of osteoarthritis. In this way, no overall bone erosion resulted, although bony nodules, so-called osteophytes, did form. We identified tumor necrosis factor-alpha (TNF) as a key inducer of DKK-1 in the mouse inflammatory arthritis model and in human rheumatoid arthritis. These results suggest that the Wnt pathway is a key regulator of joint remodeling.

A mutation in Ampd2 is associated with nephrotic syndrome and hypercholesterolemia in mice.

BACKGROUND: Previously, we identified three loci affecting HDL-cholesterol levels in a screen for ENU-induced mutations in mice and discovered two mutated genes. We sought to identify the third mutated gene and further characterize the mouse phenotype. METHODS: We engaged, DNA sequencing, gene expression profiling, western blotting, lipoprotein characterization, metabolomics assessment, histology and electron microscopy in mouse tissues. RESULTS: We identify the third gene as Ampd2, a liver isoform of AMP Deaminase (Ampd), a central component of energy and purine metabolism pathways. The causative mutation was a guanine-to-thymine transversion resulting in an A341S conversion in Ampd2. Ampd2 homozygous mutant mice exhibit a labile hypercholesterolemia phenotype, peaking around 9 weeks of age (251 mg/dL vs. wildtype control at 138 mg/dL), and was evidenced by marked increases in HDL, VLDL and LDL. In an attempt to determine the molecular connection between Ampd2 dysfunction and hypercholesterolemia, we analyzed hepatic gene expression and found the downregulation of Ldlr, Hmgcs and Insig1 and upregulation of Cyp7A1 genes. Metabolomic analysis confirmed an increase in hepatic AMP levels and a decrease in allantoin levels consistent with Ampd2 deficiency, and increases in campesterol and ß-sitosterol. Additionally, nephrotic syndrome was observed in the mutant mice, through proteinuria, kidney histology and effacement and blebbing of podocyte foot processes by electron microscopy. CONCLUSION: In summary we describe the discovery of a novel genetic mouse model of combined transient nephrotic syndrome and hypercholesterolemia, resembling the human disorder.

Inhibition of WISE preserves renal allograft function.

Wnt-modulator in surface ectoderm (WISE) is a secreted modulator of Wnt signaling expressed in the adult kidney. Activation of Wnt signaling has been observed in renal transplants developing interstitial fibrosis and tubular atrophy; however, whether WISE contributes to chronic changes is not well understood. Here, we found moderate to high expression of WISE mRNA in a rat model of renal transplantation and in kidneys from normal rats. Treatment with a neutralizing antibody against WISE improved proteinuria and graft function, which correlated with higher levels of ß-catenin protein in kidney allografts. In addition, treatment with the anti-WISE antibody reduced infiltration of CD68(+) macrophages and CD8(+) T cells, attenuated glomerular and interstitial injury, and decreased biomarkers of renal injury. This treatment reduced expression of genes involved in immune responses and in fibrogenic pathways. In summary, WISE contributes to renal dysfunction by promoting tubular atrophy and interstitial fibrosis.

Predictive potential of MRI in differentiating the predominant component in biphasic pleural mesothelioma.

PURPOSE: To assess the potential of apparent diffusion coefficient (ADC) values derived from diffusion weighted (DW) MRI preoperatively to predict the predominant histologic component among biphasic pleural mesothelioma (PM) tumors. METHODS: ADC maps were generated from DW MRI scans. Histology and predominant component of biphasic PM were confirmed following surgical resection. Statistical analyses were done with R (R Foundation for Statistical Computing, Vienna, Austria). Average ADC values corresponding to epithelioid- and sarcomatoid-predominant tumors were compared. ADC thresholding was accomplished by recursive partitioning and confirmed with ROC analysis. RESULTS: Eighty-four patients with biphasic PM's, 69 (82 %) epithelioid-predominant (BE) and 15(18 %) sarcomatoid-predominant (BS) tumors were evaluated. Thirty-eight (45 %) patients underwent extrapleural pneumonectomy (EPP), 39 (46 %) had extended pleural decortication (ePDC) and 7 (8 %) had pleural decortication (PDC). ADC values ranged between 0.696 x 10-3 to 1.921 x 10-3 mm2/s. BE tumors demonstrated significantly higher ADC values than BS tumors (p = 0.026). ADC values above 0.94 x 10-3 mm2/s were associated with a significant increase of relative risk of being in group BE over group BS (relative risk: 1.47, 95 %CI: 1.05-2.06, p = 0.027) CONCLUSION: Average ADC values of BE tumors were higher than BS tumors and the two groups can be separated by a cut off value of 0.94 X 10-3 mm2/s.