CD38 gene polymorphism rs1130169 contribution to the increased gene expression and risk of colorectal cancer (pilot study).

BACKGROUND: Genetic variations in immune signaling genes may have regulatory effect on phenotypic heterogeneity of immune cells and immune functions, hence promoting tumor growth. PURPOSE: We compared the frequencies of potentially functional CD38 gene single nucleotide polymorphisms rs1130169 (T > C) in 86 healthy controls and 90 colorectal cancer (CRC) cases to assess their association with cancer risk and CD38 gene expression. RESULTS: The association between allele C rs1130169 and CRC risk was observed. Allele C was also significantly correlated with an increased CD38 mRNA level and CD38 positive cell percentages in peripheral blood of healthy controls that could be a possible explanation for CRC risk in C allele carriers. In peripheral blood of CRC patients CD38 mRNA and serum soluble CD38 protein levels significantly differed from those in healthy controls. Calculation of the CD38 full-length and with the third exon deletion mRNA ratio in corresponding samples showed that the mRNA isoform ratio was significantly higher in CRC cases than in controls. It suggests that alternative splicing regulates elevation of CD38 full-length mRNA level in peripheral blood of CRC patients. We also have observed higher expression levels of CD38 full-length mRNA in peripheral blood of CRC patients with lymph node metastases compared to patients without metastases. CONCLUSION: This study indicated biological significance of rs1130169 variations that can alter differences in CRC risk by regulating CD38 gene expression.

SlyD-deficient Escherichia coli strains: A highway to contaminant-free protein extraction.

Binding of native bacterial protein SlyD to metal affinity matrices remains a major problem in affinity purification of His-tagged recombinant proteins from Escherichia coli cells. In this study, four novel E. coli strains that lack the expression of SlyD/SlyX, were engineered using λ-red mediated chromosomal deletion. The resultant mutant E. coli strains allow us to obtain SlyD-free proteins immediately after metal affinity chromatography, and eliminate additional purification processes. As a model protein, bispecific antibodies composed of anti-F4/80 VHH module and anti-TNF VHH module (MYSTI-2) were used. Using this protein we have shown that the SlyD/SlyX-deficient E. coli strains allow us to obtain a fully functional protein.

Treatment by serum up-conversion nanoparticles in the fluoride matrix changes the mechanism of cell death and the elasticity of the membrane.

Nanoparticles are increasingly being used for treatment and diagnostic purposes, but their effects on cells is not fully understood. Here, the interaction of fluorescent up-conversion nanoparticles (UpC-NPs) with neutrophils was investigated by imaging and measurement of membrane-cytosceletal elasticity by atomic force microscopy. It was found that UpC-NPs induce the death of neutrophils mainly by necrosis, and to a smaller extent by a novel process called 'mummification'. Necrosis occurs by gradual loss of intracellular contents and nuclei, 45-110min after exposure to UpC-NPs. Mummification is apparent as an increase in the rigidity of the neutrophils' membrane and acquisition of a characteristic bumpy shape with numerous protrusions; this structure does not change during atomic force microscopy scanning. Coating UpC-NPs with protein by incubation with serum leads to (1) formation of nanoparticle aggregates in the nm and µm size range, (2) a reduction in toxicity, (3) reduced mummification of neutrophils, and (4) no significant reduction of the elasticity of the membrane-cytoskeletal complex of neutrophils 30min after exposure to coated UpC-NPs. The study shows that serum proteins greatly curb the toxicity of nanoparticles and reveals mummification as a novel mechanism of UpC-NP-induced cell death.

Conformation-dependent evolution of copolymer sequences.

A "toy model" of molecular evolution of sequences in copolymers is proposed and implemented using a molecular-dynamics-based algorithm. The model involves coupling of conformation-dependent and sequence-dependent properties. It is shown that this model allows the realization of two main possibilities: ascending and descending branches of evolution (in terms of information content of a sequence), depending on the interaction parameters shaping the conformation of a polymer globule. The problem of adequate description of information complexity of copolymer sequences is studied. It is shown that Shannon's entropy or compressibility of a sequence gives preference to random sequences and therefore cannot be applied for this purpose. On the other hand, the Jensen-Shannon divergence measure turns out to give the description of information complexity which corresponds to our intuitive expectations. In particular, this characteristic can adequately describe two branches of evolution mentioned above, exhibiting a singularity on the boundary of these regimes.

Effect of brain injury on immunophenotype of peripheral blood lymphocytes in rats.

Strong immunosuppression occurs after severe traumatic brain injury (TBI) and most likely contributes substantially to the patient morbidity and mortality. However, the mechanisms of this immunosuppression are unknown. For the lowering of stressful factors, severe TBI was induced in anaesthetized rats. The lymphocyte subsets from 60 rats with severe TBI were analyzed using monoclonal antibody by the indirect immunofluorescence method. The blood of 30 rats without TBI was used as a control. When compared to the control group, the rats with TBI showed a remarkable reduction in the relative number of CD4(+), RT-Ia(+), Thy-1(+) and ICO-111(+) lymphocytes during the first 2 h after injury. Further reduction in the number of CD4(+) cells was determined in the rats during all the period of the experimental observation. The number of lymphocytes expressing membrane Thy-1 and ICO-111 antigens was significantly decreased 7 days after the trauma. The relative number of RT-Ia(+) lymphocytes was significantly reduced in rats with TBI during 14 days following the trauma. A significant decrease in the luminescence intensity of all the analyzed antigen-positive cells was also observed in rats with TBI. Between the 7th and the 14th days after the trauma a positive correlation between the number of Thy-1(+) PBLs and the number of RT-Ia(+) lymphocytes was determined. Similar results on lymphocyte immunophenotype were seen in patients with TBI. Thus, the cellular immune response is identical in patients and in animals with TBI. Severe brain traumatic injury leads to a reduced expression of cell surface antigens and causes a decrease in the number of antigen-positive lymphocytes and in the intensity of their luminescence.

Immunophenotype of Peripheral Blood Lymphocytes in Patients with Burns.

Severe immunosuppression occurs after large thermal burns and probably contributes substantially to patient morbidity and mortality. The mechanism of immunosuppression is much unknown. T lymphocyte subsets from 45 severely patients with burns and 35 healthy donors were analyzed using monoclonal antibodies by indirect immunofluorescence method. Compared to healthy donors, patients with burns have shown a profound reduction in relative number of CD3(+) lymphocytes during the 24-h period following injury, which was accompanied by a decrease in CD4 but not CD8 subsets. Activated lymphocytes, while determined by the expression of CD25, CD26 and CD71, were insignificantly increased in patients with burns. The expression HLA DR was insignificantly decreased on peripheral blood lymphocytes from thermally injured patients. Additionally, the significant decrease of CD95 antigen expression was observed in all patients with burns. Also, significant decrease of the luminescence intensity in all analyzed antigens was observed in patients with burns. Numerous positive correlations between CD3(+), CD8(+), CD25(+), CD26(+) and CD71(+) cells were revealed, especially during the first week after thermal trauma. We conclude that the thermal injury produces a profound relative CD3(+) and CD4(+) cell lymphopenia with signs of moderate lymphocyte activation.