DHCR24 gene expression is upregulated in melanoma metastases and associated to resistance to oxidative stress-induced apoptosis

The DHCR24 gene encoding for the 3beta-hydroxysterol delta24-reductase, an oxidoreductase involved in cholesterol biosynthesis, was isolated by subtractive hybridization as highly expressed in a short-term melanoma cell line derived from a cutaneous metastases (S/M2) compared to that obtained from the autologous primary tumor (S/P). DHCR24 (alias seladin-1, diminuto/dwarf1 homolog) has been reported to act as an antiapoptotic factor in neurons. Gene expression analysis by Northern blot confirmed that DHCR24 was 5-fold upregulated in S/M2 compared to S/P cells. High levels of DHCR24 gene expression were detected in 13/25 melanoma metastases and in 1/7 primary melanomas by real-time PCR, indicating that upregulation of this gene may occur in melanoma progression. In S/M2 cells, high DHCR24 gene expression associated with resistance to apoptosis triggered by oxidative stress induced by exposure to hydrogen peroxide. DHCR24 gene transfer was shown to protect melanoma cells from H2O2-induced cytotoxicity. Although higher cholesterol levels were shown in S/M2 cells compared to S/P cells, DHCR24 gene transfer did not increase cholesterol content. To evaluate whether DHCR24 acts as an antiapoptotic factor in melanoma metastases, the cytotoxic effect of chemotherapeutic agents was tested in DHCR24 transfectants and in the presence of a DHCR24 inhibitor, U18666A. High DHCR24 gene expression in transfectants did not result in a higher resistance to cytotoxic agents; treatment with U18666A was cytotoxic in S/P cells with a lower DHCR24 content and showed additive cytotoxic effect only when associated with H2O2 and not with cysplatin or etoposide, indicating that the DHCR24 protective effect is exerted through an oxidative stress-specific mechanism.     

β₂-glycoprotein I, the major target in antiphospholipid syndrome, is a special human complement regulator

The human plasma protein β(2)-glycoprotein I (β(2)-GPI) is the major target of autoantibodies associated with antiphospholipid syndrome. However, the biologic function of this abundant protein is still unclear. Here we identify β(2)-GPI as a complement regulator. β(2)-GPI circulates in the plasma in an inactive circular form. On surface binding, such as to apoptotic cells, β(2)-GPI changes conformation to an elongated form that acquires C3/C3b binding activities. β(2)-GPI apparently changes conformation of C3, so that the regulator factor H attaches and induces subsequent degradation by the protease factor I. β(2)-GPI also mediates further cleavage of C3/C3b compared with factor H alone. Our data provide important insights into innate immune regulation by plasma protein β(2)-GPI, which may be exploited in the prevention and therapy of autoimmune disease antiphospholipid syndrome.     

Clonal drift demonstrates unexpected dynamics of the T-cell repertoire in T-large granular lymphocyte leukemia

T-cell large granular lymphocyte leukemia (T-LGLL) is characterized by chronic lymphoproliferation of cytotoxic T lymphocytes (CTLs) and is associated with lineage-restricted cytopenias. Introduction of T-cell receptor (TCR) variable β-chain (Vβ) monoclonal antibodies has facilitated identification and enumeration of clonal CTLs by flow cytometry. A highly skewed TCR Vβ repertoire identified by flow cytometry is strongly associated with monoclonal CDR3 regions by quantitative sequencing and positive TCRγ rearrangement assays. Therefore, Vβ expansions can serve as surrogate markers of CTL clonality to assess clonal kinetics in T-LGLL. We analyzed the TCR repertoire in 143 patients, 71 of which were available for serial measurements over 6 to 96 months. Although the majority (38/71, 54%) maintained a consistent monoclonal expansion, many (26/71, 37%) unexpectedly displayed a change in the dominant clone, whereby the original CTL clone contracted and another emerged as demonstrated by Vβ typing. Our results demonstrate that the T-cell repertoire is more dynamic in T-LGLL than recognized previously, illustrating the heterogeneity of disorders under this categorization.     

Physiological and pharmacological properties of a modified Brooke ileostomy: justification for retaining the most distal ileum

Purpose

The traditional Brooke ileostomy removed the last 8–15 cm of the ileum due to concern of occurrence of terminal ileal Crohn’s disease, vide infra the ileocolic sphincter was removed. Retaining all the terminal ileum has the potential of retaining the ileocolic sphincter. Our aim was to investigate whether a high-pressure zone existed within the last few centimetres of the ileum and its response to pharmacological stimuli.

Methods

A balloon manometry catheter was introduced into the stoma of 16 patients who had formation of an end ileostomy (ileocolic sphincter retained, ICS). Recordings were made at 1 cm intervals from the meatus in order to identify the maximum intra-luminal resting and intra-abdominal pressure. At the point of maximum resting pressure, the response to phenylephrine (10 % gel) and glyceryl trinitrate (GTN) (0.2 % paste) was recorded. Results were recorded using an Ohmeda Oestiva 5 manometry system (in millimeter of mercury) and data were analysed using ANOVA. Results were compared with 13 historical controls (ileocolic sphincter removed).

Results

There was no significant difference in resting intra-abdominal pressure between the two groups (historical controls 8.5?±?3.0 mmHg; ICS 9.0?±?3.2 mmHg), p?=?NS. The maximum resting intra-luminal pressure in ICS patients exceeded historical controls 16?±?2.9 vs 10.0?±?2.5 mmHg, p?<?0.001. In ICS patients, phenylephrine increased the resting pressure to 26.0?±?3.5 mmHg, p?<?0.001. In historical controls, the pressure remained unchanged, 12?±?4.7 mmHg, p?=?NS. Subsequent addition of GTN to both groups lowered maximum intra-luminal pressure to pre-study values, 10?±?4.2 mmHg (ICS) and 7?±?3.5 mmHg (controls), p?=?NS.

Conclusion

Retention of the ileocolic sphincter in a modified Brooke ileostomy preserves a physiological high-pressure zone, the properties of which can be modified by pharmacological agents.     

Mutational Signature Analysis Reveals NTHL1 Deficiency to Cause a Multi-tumor Phenotype

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Inhibition of the growth of pre-established subcutaneous tumor nodules of human prostate cancer cells by single injection of the recombinant adenovirus p53 expression vector

We have previously described potent growth-inhibitory effect of a recombinant adenovirus expressing wild type p53 (AdWTp53) in metastatic prostate cancer cells via activation of cellular p53 pathways. We have extended these observations to analyze the effects of AdWTp53 on primary cultures of radical prostatectomy specimens (RPS) and have also evaluated the gene therapeutic potential of the AdWTp53 in a nude mice model. Infection of primary cultures of prostate cancer specimens resulted in about 80% cell growth inhibition in comparison with cultures treated with control adenovirus dl312. Single injection of AdWTp53 into pre-established tumor nodules of DU145 prostate cancer cells suppressed tumor growth significantly (p = 0.0407) as determined by comparison of tumor volumes of the AdWTp53-treated vs. control vector (dl312) or PBS-treated groups. Moreover, there was no significant difference in tumor growth inhibition between single vs. multiple injections of AdWTp53. Our observations support the potential of AdWTp53 for gene therapy of prostate cancer. Int. J. Cancer 71:377-382, 1997. © 1997 Wiley-Liss Inc.