Establishment and characterization of novel patient-derived osteosarcoma xenograft and cell line

Osteosarcoma is an aggressive mesenchymal malignancy of the bone. Patient-derived models are essential tools for elucidating the molecular mechanisms associated with poor prognosis and the development of novel anticancer drugs. This study described the establishment of a patient-derived cancer model of osteosarcoma. Primary osteosarcoma tumor tissues were obtained from an osteosarcoma patient and inoculated in the skin of immunodeficient mice, followed by transplantation to other mice upon growth. Cells were maintained in monolayer cultures, and the capability of spheroid formation was assessed by seeding the cells on culture dishes. The invasion ability of cells was monitored by Matrigel assay, and genomic and proteomic backgrounds were examined by mass spectrometry. A cell line was established from patient-derived tumors and showed similar histology to that of the primary tumor tissue. Additionally, these cells formed spheroids on low-attachment tissue-culture dishes and exhibited invasive capabilities, and we confirmed that the genomic backgrounds were similar between patient-derived xenograft tumors and the cell line. Furthermore, the proteome of the patient-derived tumors and the cells exhibited similar, but not identical, patterns to that of the original tumor tissue. Our results indicated that this patient-derived xenograft model and cell line would be useful resources for osteosarcoma research.     

Preclinical and clinical studies on cancer-associated cachexia

Background

Cancer cachexia is the wasting condition that is often seen in advanced stage cancer patients. This wasting is largely attributable to a systemic and progressive loss of skeletal muscle mass that greatly hinders performance of normal daily activities, resulting in reduced quality of life. Moreover, it negatively influences the prognosis of cancer patients. A general consensus in the field is that the loss of muscle mass is due both to an increase in protein degradation and a decrease in protein synthesis. Recent studies using preclinical models for studying cachexia have been useful in identifying the contribution of inflammatory cytokines (e.g. tumor necrosis factor-α and Interleukin-6), and myostatin receptors (e.g. the type IIB activin receptor) to cachexia development, and have led to several clinical trials. However, many questions remain about the molecular mechanisms thought to play a role in the development of cachexia.

Methods

We conducted a literature search using search engines, such as PubMed and Google Scholar to identify publications within the cancer cachexia field.

Results

We summarized our current knowledge of: 1) the driving mechanisms of cancer cachexia, 2) the preclinical models available for studying the condition, and 3) the findings of recent clinical trials.

Conclusion

Cancer cachexia is a complex and variable condition that currently has no standard effective therapeutic treatment. Further studies are desperately needed to better understand this condition and develop effective combination treatments for patients.

     

Virulence of Yersinia pseudotuberculosis isolated from pork and from the throats of swine

Yersinia pseudotuberculosis was isolated from retail pork and from healthy swine throats. These wild-type strains and their representative cured isogenic strains were tested for the presence of plasmids and several virulence factors, and these characteristics were compared with those of virulent strains from humans. Two pork isolates (serotype IVB) and four swine isolates (serotypes IIB, IIC, III, and IVB) harbored a 42- to 48-megadalton plasmid which had similar fragmentation patterns resulting from digestion with restriction endonuclease. These six strains were lethal for mice via oral challenge and were positive in autoagglutination and calcium dependency tests. They also invaded HeLa cells and induced cytotoxicity. Histopathological examination and indirect fluorescent-antibody staining provided definite evidence of the pathogenicity of these strains when tissue sections from orally infected mice were used. The virulence factors of wild-type pork and swine isolates with the 42- to 48-megadalton plasmid were identical to those of two human isolates (serotypes IVB and VB). Hence, these pork and swine isolates should be considered potentially pathogenic for humans. The finding suggests that retail pork and swine may play an important role in the epidemiology of human infections caused by Y. pseudotuberculosis.     

Chlorination of harman and norharman with sodium hypochlorite and co-mutagenicity of the chlorinated products

Harman and norharman are widely distributed in the environment and consequently contaminate in domestic waste-water. It has been reported that they have co-mutagenic activity in the presence of non- mutagenic aromatic amines such as aniline and o-toluidine with S9 mix. When these β-carbolines were treated with sodium hypochiorite under mild conditions, chlorinated derivatives were produced. Among them, 6-chloroharman and 6-chloronorharman showed much more potent co-mutagenic activities than harman and norharman in the presence of o-toluidine toward Salmonella typhimurium TA98 with S9 mix. These results suggest that the chlorination of harman and norharman occurs during disinfection at the sewage plant to produce potent co-mutagens that contaminate river water.     

Nitrogen budget and gaseous nitrogen loss in a tropical agricultural watershed

Although agricultural systems in tropical monsoon Asia play a central role in the global nitrogen (N) cycle, details of the N cycle in this region on a watershed scale remain unclear. This study quantified the N budget in a tropical watershed of 221 km² on Java Island, where paddy fields cover 28% of the land, by conducting field surveys. The amount of net biochemical gaseous N loss to the atmosphere (X _GB ), which is generally difficult to determine, was calculated as the residual of the N balance. Assuming that NH₃ volatilization balances deposition, and hence subtracting NH₄–N from the N import with atmospheric deposition, the average total import and export of N per year was found to be 46.5 kg ha⁻¹ year⁻¹ over the watershed. Of this, 71% was imported as fertilizer (M _F ) and 29% with atmospheric deposition (M _AD ). On the export side, 42% was lost as X _GB , 37% with incineration of rice residues and wood fuel (X _GI ), 13% with river discharge (X _D ) and 9% with rice surplus export (X _R ). A large portion of X _GB , and consequently, a small portion of X _D could be explained by the high rate of denitrification resulting from the high temperature and humid climate, and are thought to be common features of tropical watersheds where paddy fields are found.     

Pluvial Drainage Patterns and Holocene Desiccation Influenced the Genetic Architecture of Relict Dace,Relictus solitarius (Teleostei: Cyprinidae)

Changing drainage patterns have played a significant role in the evolution of western North American aquatic taxa. Relict dace, Relictus solitarius, is a Great Basin endemic cyprinid with a native range that is restricted to four valleys in eastern Nevada. Relictus solitarius now occupies spring systems that are the remnants of Pleistocene-era pluvial lakes, although it may have occurred in the area for much longer. Here we use mitochondrial DNA sequence data to assess range-wide genetic diversity of R. solitarius, and to estimate divergence times to determine whether pluvial drainages played an important role in shaping intraspecific genetic diversity. Genetic diversification within R. solitarius began during the early to mid-Pleistocene, separating populations within two sets of valleys (Butte/Ruby and Goshute/Steptoe). Additional diversification in each of the two sets of valleys occurred more recently, in the mid- to late-Pleistocene. Holocene desiccation has further isolated populations, and each population sampled contains unique mtDNA haplotypes. Pluvial drainage patterns did contribute to the genetic structure observed within R. solitarius, but most of the intraspecific diversification does not appear to be associated with the Last Glacial Maximum. Holocene desiccation has also contributed to the observed genetic structure. The relict dace populations we sampled are all unique, and we recommend that future management efforts should strive to preserve as much of the genetic diversity as possible.     

Ring-fused pyrazole derivatives as potent inhibitors of lymphocyte-specific kinase (Lck): Structure,synthesis, and SAR

We have identified a novel series of ring-fused pyrazole derivatives as lymphocyte-specific kinase (Lck) inhibitors. The most potent analogs exhibited good enzyme inhibitory activity (IC₅₀s <1 nM) as well as excellent cellular activity against mixed lymphocyte reaction (MLR) (IC₅₀s <1 nM).     

Efficient sequential gene regulation via FLP-and Cre-recombinase using adenovirus vector in mammalian cells including mouse ES cells

Site-specific recombinase is widely applied for the regulation of gene expression because its regulatory action is strict and efficient. However, each system can mediate regulation of only one gene at a time. Here, we demonstrate efficient "sequential" gene regulation using Cre-and FLP-expressing recombinant adenovirus (rAd) in two different monitor cell lines, for regulation of one gene (OFF-ON-OFF) and for two genes (ON-OFF and OFF-ON, independently). Generally, serial use of Cre-and FLP-expressing rAd tends to cause significant cytotoxicity, but we here described optimum dose of the rAds for serial regulation. We also established an efficient method of rAd infection to mouse ES cell lines after removing feeder cells, showing that this system is useful for removal of FRT-flanked drug-resistance gene cassette from recombinant ES cells prior to introduction of ES cells into blastocytes for chimeric mice production. Because our sequential gene-regulation system offers efficient purpose-gene regulation and strict OFF-regulation, it is potentially valuable for elucidating not only novel gene functions using cDNA microarray analysis but also for "gene switching" in development and regeneration research.     

Treatment of arthritis with a selective inhibitor of c-Fos/activator protein-1

To inhibit arthritis upstream of inflammatory cytokine release and matrix metalloproteinase (MMP) action, we designed de novo a small-molecule inhibitor of c-Fos/activator protein-1 (AP-1) using three-dimensional (3D) pharmacophore modeling. This model was based on the 3D structure of the basic region-leucine zipper domain of AP-1-DNA complex. Administration of this inhibitor prevented type II collagen-induced arthritis from day 21, before the onset of arthritis, or from day 27, resolved arthritis after its onset. Suppression of disease was accomplished by reducing the amounts of inflammatory cytokines and MMPs in vivo in sera and joints and in vitro in synovial cell and chondrocyte cultures. The primary action of this molecule was the inhibition of matrix-degrading MMPs and inflammatory cytokines including interleukin 1beta; this molecule also synergized with anti-tumor necrosis factor alpha to inhibit arthritis. Thus, selective inhibition of c-Fos/AP-1 resolves arthritis in a preclinical model of the disease.     

The role of CXCR7/RDC1 as a chemokine receptor for CXCL12/SDF-1 in prostate cancer

Several reports have recently documented that CXCR7/RDC1 functions as a chemokine receptor for SDF-1/CXCL12, which regulates a spectrum of normal and pathological processes. In this study, the role of CXCR7/RDC1 in prostate cancer (PCa) was explored. Staining of high density tissue microarrays demonstrates that the levels of CXCR7/RDC1 expression increase as the tumors become more aggressive. In vitro and in vivo studies with PCa cell lines suggest that alterations in CXCR7/RDC1 expression are associated with enhanced adhesive and invasive activities in addition to a survival advantage. In addition, it was observed that CXCR7/RDC1 levels are regulated by CXCR4. Among the potential downstream targets of CXCR7/RDC1 are CD44 and cadherin-11, which are likely to contribute to the invasiveness of PCa cells. CXCR7/RDC1 also regulates the expression of the proangiogenic factors interleukin-8 or vascular endothelial growth factor, which are likely to participate in the regulation of tumor angiogenesis. Finally, we found that signaling by CXCR7/RDC1 activates AKT pathways. Together, these data demonstrate a role for CXCR7/RDC1 in PCa metastasis and progression and suggest potential targets for therapeutic intervention.