Enhancing effects of monocytes on modulation of a lymphocyte membrane antigen

Redistribution, or modulation, of some cell surface antigens occurs in the presence of specific antibody. The phenomenon of antigenic modulation may therefore affect the use of antibodies as therapeutic agents. This study was undertaken to investigate modulation of the 65,000 dalton T65 antigen, present on normal and malignant T cells and some malignant B cells, which is recognized by the monoclonal antibody T101. To induce cell surface antigenic modulation, normal or leukemic lymphoid cells were cultured in the presence of monoclonal antibody T101 for 3-hr periods. Removal of monocytes from mononuclear cell preparations resulted in significantly lower degrees of T65 antigenic modulation. The degree of antigenic modulation could be increased by adding monocytes back to monocyte-depleted lymphocyte suspensions. Furthermore, maximal modulation occurred in the presence of monocytes at T101 concentrations that were 3 logs lower than in the absence of monocytes. The enhancing effect of monocytes was dependent on the Fc portion of the T101 antibody molecule, and presumably was mediated by cross-linking of antigen-antibody complexes on the surface membrane of the modulating cell by Fc receptors present on monocytes. Further experiments performed to examine the characteristics of this enhancement of antigenic modulation by monocytes indicated that autologous as well as allogeneic monocytes were effective, indicating that the enhancing phenomenon was not dependent upon recognition of major histocompatibility antigens. Viable monocytes were required, but pretreatment of monocytes with sodium azide to inhibit energy production, or indomethacin to inhibit prostaglandin synthesis had no effect on this phenomenon. Polymorphonuclear leukocytes did not mediate similar enhancement, although monocytic and myeloid cell lines U937, THP-1, and HL-60 did. Spent culture medium from modulated cultures and preparations containing IL 1 activity did not enhance modulation of the T65 surface antigen on lymphocytes, suggesting that direct contact between lymphocytes and monocytes is required to mediate the effect. The finding that leukemic cells from patients with CLL undergo modulation of the T65 antigen to a much lower degree in vitro than observed in vivo, and that this difference can be overcome by the addition of monocytes, suggests that monocytes or the reticuloendothelial system may augment antigenic modulation in vivo.     

Substituted hydrophobic and hydrophilic residues at methionine-68 influence the chaperone-like function of αB-crystallin

Amino acid residues 57–69 in B-crystallin have been implicated as a target protein binding site. Moreover, a direct correlation between the extent of -crystallin hydrophobicity and chaperone-like activity has been demonstrated. The purpose of this study was to mutate a moderately hydrophobic residue Met-68 (M-68) in the above region to strongly hydrophobic and hydrophilic residues and show whether chaperoning ability is affected with or without structural changes. Mutation of M-68 to Val, Ile or Thr did not result in significant changes in molecular mass and secondary and tertiary structures. However, the Val and Ile mutants showed significant improvement and the Thr mutant showed substantial loss in chaperone activity. Differences in chaperone function in the absence of any structural changes confirmed that the hydrophobicity or hydrophilicity of the substituted amino acid in the putative target protein binding site was the only contributing factor.     

Detection of the intracellular ras p21 oncogene product by flow cytometry

Rapid identification of the expression of oncogene products in specific cell types could potentially be useful in the diagnosis and treatment of human malignancy. We have now observed that through the use of lysolecithin permeabilization and fluorescence-activated flow cytometry, cells expressing high levels of the v-Ha-ras oncogene product, p21, can readily be distinguished from the nontransformed parent cells in a rapid and quantitative manner.     

Spatial distribution of pollen-induced symptoms within a large metropolitan area—Berlin,Germany

Large spatial differences in the distribution of three allergologically relevant pollen types for Central Europe—birch, grass, and mugwort—are revealed within a large metropolitan area—Berlin, Germany. The purpose of the study is an examination of the hypothesis that these different pollen exposure conditions can cause different degrees of pollen-induced symptoms within the city. Pollen data from 14 gravimetric traps and one volumetric trap in Berlin and anonymously reported pollen-induced symptom data from the online-based self-documentation tool “Patient’s Hayfever Diary” (PHD) are used for the analysis of temporal and spatial variations of the severity of the overall total symptoms. Geographically localised symptom data are linked to the nearest pollen trap. Statistical analysis is performed using Kendall’s Tau-b. Higher amounts of monitored birch and grass pollen in the peripheral areas of Berlin induce stronger symptoms in PHD users located within suburbs than those located in the city centre. There is no statistical relationship between the varying presence of mugwort pollen in the air and the severity of symptoms. Spatial differences in the pollen-induced symptom severity within a large city coinciding with spatial differences in birch and grass pollen depositions are shown for the first time. Therefore, pollen data from a single trap may not provide an appropriate explanation for differences in pollen-induced symptoms across the city. More detailed and reliable information about the exposure to allergenic pollen can be addressed by installing further traps in order to improve the knowledge about pollen exposure within cities.     

Peripheral non-viral MIDGE vector-driven delivery of β-endorphin in inflammatory pain

Background

TRPA1 has been implicated in both chemo- and mechanosensation. Recent work demonstrates that inhibiting TRPA1 function reduces mechanical hypersensitivity produced by inflammation. Furthermore, a broad range of chemical irritants require functional TRPA1 to exert their effects. In this study we use the ex-vivo skin-nerve preparation to directly determine the contribution of TRPA1 to mechanical- and chemical-evoked responses at the level of the primary afferent terminal.

Results

Acute application of HC-030031, a selective TRPA1 antagonist, inhibited all formalin responses in rat C fibers but had no effect on TRPV1 function, assessed by capsaicin responsiveness. Genetic ablation experiments corroborated the pharmacological findings as C fibers from wild type mice responded to both formalin and capsaicin, but fibers from their TRPA1-deficient littermates responded only to capsaicin. HC-030031 markedly reduced the mechanically-evoked action potential firing in rat and wild type mouse C fibers, particularly at high-intensity forces, but had no effect on the mechanical responsiveness of Aδ fiber nociceptors. Furthermore, HC-030031 had no effect on mechanically-evoked firing in C fibers from TRPA1-deficient mice, indicating that HC-030031 inhibits mechanically-evoked firing via a TRPA1-dependent mechanism.

Conclusion

Our data show that acute pharmacological blockade of TRPA1 at the cutaneous receptive field inhibits formalin-evoked activation and markedly reduces mechanically-evoked action potential firing in C fibers. Thus, functional TRPA1 at sensory afferent terminals in skin is required for their responsiveness to both noxious chemical and mechanical stimuli.     

HCPTPA, a protein tyrosine phosphatase that regulates vascular endothelial growth factor receptor-mediated signal transduction and biological activity

Angiogenesis is a tightly controlled process in which signaling by the receptors for vascular endothelial growth factor (VEGF) plays a key role. In order to define signaling pathways downstream of VEGF receptors (VEGFR), the kinase domain of VEGFR2 (Flk-1) was used as a bait to screen a human fetal heart library in the yeast two-hybrid system. One of the signaling molecules identified in this effort was HCPTPA, a low molecular weight, cytoplasmic protein tyrosine phosphatase. Although HCPTPA possesses no identifiable phosphotyrosine binding domains (i.e. SH2 or phosphotyrosine binding domains), it bound specifically to active, autophosphorylated VEGFR2 but not to a mutated, kinase-inactive VEGFR2. Recombinant VEGFR2 and endogenous VEGFR2 were substrates for recombinant HCPTPA, and HCPTPA was co-expressed with VEGFR2 in endothelial cell lines, suggesting that HCPTPA may be a negative regulator of VEGFR2 signal transduction. To pursue this possibility, an adenovirus directing the expression of HCPTPA was constructed. When used to infect cultured endothelial cells, this adenovirus directed high level expression of HCPTPA that resulted in impairment of VEGF-mediated VEGFR2 autophosphorylation and mitogen-activated protein kinase activation. Adenovirus-mediated overexpression of HCPTPA also inhibited VEGF-induced cellular responses (endothelial cell migration and proliferation) and inhibited angiogenesis in the rat aortic ring assay. Taken together, these findings indicate that HCPTPA may be an important regulator of VEGF-mediated signaling and biological activity. Potential interactions with other signaling pathways and possible therapeutic implications are discussed.     

Anaerobic reduction of fumarate in the body wall musculature ofArenicola marina (Polychaeta)

Summary The reduction of fumarate, which is a characteristic feature of anoxic catabolism of some invertebrates, was investigated in mitochondria or mitochondrial fragments prepared from the body wall musculature of the lug-wormArenicola marina (Annelida, Polychaeta).A coupling of the reduction of fumarate to succinate to the oxidation of NADH was demonstrated.The pathway of hydrogen transfer from NADH to fumarate was studied by using specific inhibitors of the respiratory chain. From the results it is concluded that parts of the respiratory chain are involved.During anaerobiosis mitochondria formed succinate at a high rate from malate which had been added as substrate. The formation of succinate is coupled to oxidative phosphorylation. The ratio ATP-production/formation of succinate was found to be 0.6 to 0.8.Succinate formation from malate is inhibited by arsenite and monofluoroacetate.TheK _m for fumarate of the fumarate reductase inArenicola body wall musculature is 2.5×10^–5 M.Abbreviations Ap₅A P¹,P⁵-di(adenosine-5-)pentaphosphate - APAD acetylpyridine adenine dinucleotide - DNP 2,4-dinitrophenol - fw fresh weight (of body wall musculature) - NaFAc sodiummonofluoroacetic acid - PCA perchloric acid - PEP phosphoenolpyruvate Supported by Deutsche Forschungsgemeinschaft (Ze 40/13, Ze 40/14 and Gr 456/5)     

Regeneration of T cell subpopulations after bone marrow transplantation: cytomegalovirus infection and lymphoid subset imbalance

Immune reconstitution was studied serially by using T lymphocyte cell surface differentiation antigens in 37 individuals receiving bone marrow transplants. Antigen expression was assessed by immunofluorescence analysis using monoclonal antibodies to T lymphocytes including Leu-3, which defines a T lymphocyte subpopulation in healthy individuals with T helper or inducer activity (L3+), and Leu-2, which defines a T lymphocyte subpopulation in healthy individuals with T helper or inducer activity (L3+), and Leu-2, which defines a T lymphocyte subpopulation with suppressor or cytotoxic activity (L2+). These studies demonstrated that the L2+ subpopulation regenerated more rapidly after transplant than did the L3+ subpopulation. Imbalances between these two T lymphocyte subpopulations, indicated by a decreased L3/L2 ratio, persisted for periods up to 12 mo post-transplant. Expression of a cell surface antigen associated with immature lymphocytes (OKT-10), and of HLA-DR (Ia-like) antigens was markedly increased during the post-transplant period. HLA-DR antigen expression did not appear related to immune activation in that increased reactivity was not detected with a monoclonal antibody (anti-TAC) specific for activated T cells. These observations in bone marrow transplant recipients and other disorders characterized by lymphoid restoration or immaturity indicate that inversion of the normal L3/L2 ratio and increased expression of OKT-10 and HLA-DR antigens may be features of a regenerating immune system. Furthermore, serial observation of individual patients indicated that infection with cytomegalovirus was associated with a progressive decrease in the L3/L2 ratio.     

Google Trends reflect allergic rhinitis symptoms related to birch and grass pollen seasons

Google Trends (GT) describes the variation of the relevant interest of internet searches toward medical conditions and related symptoms. Allergic rhinitis symptom levels result from the intensity of exposure to aeroallergens in combination with relevant medication use. We analyze data from Germany to examine the relationship between hay fever-related Google search terms, symptom levels, medication use, and pollen count levels. For doing so, we also employ the new definitions on pollen season and peak pollen period start and end as proposed by the European Academy of Allergy and Clinical Immunology in a recently published position paper. We extract GT data for a number of search terms related to allergic rhinitis for Germany. We use total nasal symptom and mediation scores as reported by patients via a patient hay fever diary in the Berlin and Brandenburg areas in Germany for 3 years (2014–2016), accompanied by pollen data. Then a Pearson and Spearman correlation analysis is performed between symptom data and GT data. A graphical analysis is conducted, and the identification of pollen season and peak pollen periods is done based on the EAACI criteria. The analysis reveals that GT data are highly correlated with symptom levels and follow peak pollen period start–end, concerning grass and birch pollen-induced allergic rhinitis symptoms. GT data can be used as a proxy for the identification of the onset and variation of nasal symptom and medication score for allergic rhinitis sufferers.