Kappa and lambda light chain proteins--clinical and prognostic significance in patients with multiple myeloma

In 76 patients with multiple myeloma an independent or combined light chain production at a ratio of kappa (kappa) to lambda (lambda) chains of 43:33 was proved. Two groups of patients were formed depending on the type of the light chain production. They were compared by a number of biological, clinical and biochemical parameters, which demonstrate the frequency and pathogenetic participation of the two light chains in the main syndromes of the disease. The therapeutic response and prognostic value were also estimated. Bence Jones (lambda) chains prevail in men, in the III clinical stage of the disease, in patients with tubular proteinuria and in syndromes indicating an advanced stage of evolution of the main process, in non-reversible azotaemia, hypercalcaemia and hypoalbuminemia. The survival rate in thus group of patients is an average of 7 months shorter in comparison with the BJ (kappa) group (the difference is non-significant). The median survival of patients with BJ (kappa) is 30 months and 21 months for BJ (lambda).     

Molecular analysis of human immunoglobulin heavy chain variable genes (IgVH) in normal and malignant B cells

The diagnostic value of monitoring human cytomegalovirus (HCMV) late pp67 mRNA expression by nucleic acid sequence-based amplification (NASBA) after renal-allograft transplantation was evaluated. RNAs were isolated from 489 whole-blood specimens of 42 patients for the specific amplification of the late pp67 (UL65) mRNA. NASBA results were compared to results from the pp65 antigenemia assay, virus isolation by cell culture, and serology. The sensitivity value for NASBA proved to be higher than that for the antigenemia assay (50 versus 35%) for the detection of HCMV infection, while the sensitivity values of cell culture and NASBA were comparable (54 and 50%, respectively). NASBA detected the onset of HCMV infection simultaneously with cell culture and the antigenemia assay. Both the antigenemia assay and NASBA are very specific (100%) and highly predictive (100%) for the onset of HCMV infection. Antiviral therapy with ganciclovir resulted in negative results for cell culture, the antigenemia assay, and NASBA. In conclusion, monitoring HCMV pp67 mRNA expression by NASBA is a highly specific method for the detection of HCMV infection in renal-allograft recipients and is more sensitive than the antigenemia assay. Furthermore, NASBA can be used to monitor the progression of HCMV infections and the effect of antiviral therapy on viral activity.     

Cellular and molecular factors that regulate the differentiation and apoptosis of germinal center B cells. Anti-Ig down-regulates Fas expression of CD40 ligand-stimulated germinal center B cells and inhibits Fas-mediated apoptosis

To investigate the molecular and cellular mechanisms underlying the selection, differentiation, and apoptosis of germinal center (GC) B cells, we have established a culture system containing a follicular dendritic cell (FDC) line, HK. The mAb, 3C8, which is specific to HK cells and recognizes dendritic network in the GC, was developed and provided additional evidence that HK cells are related to FDC by sharing a unique surface Ag. The roles for CD40 ligand (CD40L) and T cell-derived cytokines in the differentiation of GC B cells were investigated in our culture system. We show that there are two distinct stages of GC B cell differentiation. In the early stage, GC B cells undergo spontaneous apoptosis unless they are stimulated by CD40L. In the secondary stage, IL-10 directs GC B cell differentiation toward the generation of plasma cells, while the absence of IL-10 stimulation leads to the generation of memory B cells. The major function of CD40L was found in the enhancement of cell recovery and the augmentation of memory B cell generation. Although GC B cells are Fas+, GC B cells are at first resistant to, but then become sensitive to, anti-Fas killing after 24 h in culture with CD40L, which coincides with the gradual increase in Fas expression on GC B cells. Furthermore, anti-Ig down-regulated Fas expression on CD40L-stimulated GC B cells, suggesting that Ag receptor engagement down-regulates Fas expression and prevents Fas-mediated apoptosis of GC B cells. Our data imply that GC T cells have an important role in the differentiation and apoptosis of GC B cells. GC T cells expressing both CD40L and Fas ligand have a dual function on GC B cells, helper or killer, depending on the status of target B cells. In the early stage, GC T cells stimulate the extensive proliferation of GC B cells, ensuring a large repertoire of B cells for selection. In the later stage, GC T cells kill B cells via Fas-Fas ligand interactions unless GC B cells are positively selected by Ags present on FDC.     

Dissociation of left ventricular hypertrophy, beta-myosin heavy chain gene expression, and myosin isoform switch in rats after ascending aortic stenosis

The regulation of angiotensin II (Ang II) receptors and Ang II-induced modulation of intracellular Ca2+ concentration in cardiac cells from hearts of experimentally induced hypertensive deoxycorticosterone acetate (DOCA)-salt and control unilaterally nephrectomized (Uni-Nx) Sprague-Dawley rats was assessed. Ang II receptor density and intracellular Ca2+ concentration measurements were examined in adult ventricular myocytes and fibroblasts by radioligand binding assay and digital imaging using fura 2 methodology, respectively. Four-week DOCA-salt treatment induced hypertension associated with cardiac hypertrophy. Ang II binding studies demonstrated that adult ventricular myocytes and fibroblasts possess mainly the AT1 subtype receptor. Moreover, DOCA-salt hypertension was associated with a 1.8-fold increase in Ang II-specific binding compared with myocytes from Uni-Nx control rats. Intracellular Ca2+ responses induced by increasing Ang II concentrations (10[-12] to 10[-4] mol/L) were significantly enhanced in cardiomyocytes from DOCA-salt rats. The effects of Ang II on intracellular Ca2+ spike frequency were unaltered in cardiomyocytes from DOCA-salt-hypertensive rats. The density of AT1 subtype receptors was not modified in ventricular fibroblasts after DOCA-salt treatment. Ang II increased intracellular Ca2+ concentration similarly in ventricular fibroblasts from normal and hypertensive rats. In conclusion, DOCA-salt hypertension is characterized by an increased AT1 receptor density and intracellular calcium responses in ventricular myocytes, whereas in ventricular fibroblasts the AT1 receptor status is unaltered. These findings report for the first time the cardiac cell-specific implication of Ang II and the intracellular calcium signaling pathway stimulated by the AT1 receptor in cardiac hypertrophy in DOCA-salt-hypertensive rats.     

The murine Fhit locus: isolation, characterization, and expression in normal and tumor cells

The murine Fhit locus maps near the centromere nu proximal Ptprg locus on mouse chromosome 14. The cDNA sequence and structure are similar to those of the human gene, with exons 5-9 encoding the protein. The predominant mRNA in the tissues and cell lines tested was an alternatively spliced form missing exon 3. Most murine cell lines tested, including lines established from normal mouse embryos and tumors, expressed very low or undetectable levels of Fhit mRNA. Most normal mouse tissues expressed wild-type Fhit mRNA, whereas approximately 40% of murine lung carcinomas expressed wild-type and aberrant Fhit RT-PCR products that lacked various exons. Several tumorigenic mouse cell lines exhibited homozygous deletions of Fhit exons. We conclude that the murine Fhit gene, like its human counterpart, is a target of alterations involved in murine carcinogenesis.     

Cysteine protease CPP32, but not Ich1-L, is expressed in germinal center B cells and their neoplastic counterparts

Ich-1/Nedd2 and CPP32/YAMA are cysteine proteases related to interleukin 1-beta-converting enzyme (ICE), which act as apoptosis effectors. Both molecules are expressed in T- and B-cell lines. The authors investigated their in vivo cellular distribution in normal and neoplastic human lymphoid tissues. Sixty-eight representative non-Hodgkin's lymphomas (NHL) and Hodgkin's disease (HD) samples, normal lymphoid organs, and nonlymphoid tumors were analyzed by immunohistochemistry (IHC). CPP32 expression in benign tissues was restricted to germinal center B cells, plasma cells, and a few interfollicular immunoblasts. All follicular NHLs and most diffuse large cell NHLs were CPP32 positive. Among T-cell NHLs, CPP32 expression was mainly observed in anaplastic large cell NHLs, whereas the other subtypes were less frequently positive. In contrast, lymphoid organs displayed only weak Ich1-L expression, located in sinusal histiocytes and thymic epithelial cells. Lymphomas were Ich1-L negative, except for T-cell-rich B-cell NHLs, and about half of the HD samples, in which Reed-Sternberg cells (RSC) were usually Ich1-L positive/CPP32 negative. Extralymphoid Ich1-L reactivity was found in particular organs like the kidney and various tumors. Western blot analysis confirmed the specificity of immunostaining. Neither CPP32 nor Ich1-L expression were correlated with intratumoral DNA fragmentation, as determined by the TUNEL assay. Altogether, these results indicate that CPP32 is preferentially expressed in germinal centers and thus could be involved in B-cell maturation. The differential expression of CPP32 and Ich1-L suggests that cysteine proteases differ in substrate specificities and carry out functions unrelated to apoptosis.     

The polymorphism of the immunoglobulin heavy chain switch region gene in IgA nephropathy, Henoch-Sch?nlein nephritis and idiopathic nephrotic syndrome

It is well known genetic predisposition may play an important role in the pathogenesis of renal diseases. Recently, there has been some controversy about the possible role of the polymorphism of the immunoglobulin heavy chain switch region gene. We have studied this gene by SstI restriction fragment length polymorphisms using DNA from 41 children with IgA nephropathy, 44 with Henoch-Sch?nlein nephritis and 60 with idiopathic nephrotic syndrome. There was no association of specific genotype with these diseases, in contrast to previous reports. These results are probably due to the differing genetic background of Japanese and Caucasoid patients as far as the switch region is concerned; no switch region genotype constitutes a genetic risk factor for the Japanese in these diseases.     

Differential expression of human ferritin H chain gene in immortal human breast epithelial MCF-10F cells

This study investigated fractures of the cement layer between a crown and a die. Model crowns were luted on dies with zinc phosphate cement and various magnitudes of load were applied. Surface strain on the crown was measured using strain gauges. Seal was evaluated using dye penetration and tensile tests. Results showed that cement fracture affected surface strain behavior on the crown and was detected using the surface strain measurement. The dye penetration test and the tensile test could not be used to detect the cement fracture. It is suggested that the three-dimensional relative positioning between the crown and the die affected the development of the cement fracture.     

Tissue inhibitor of metalloproteinase (TIMP)-1 induces differentiation and an antiapoptotic phenotype in germinal center B cells

Tissue inhibitors of metalloproteinases (TIMPs) have been shown to be multifunctional factors. Contrasting with their enzyme-inhibitory activity, TIMPs also promote cell growth. Previously, we have reported an enhanced expression of TIMP-1 by normal reactive B cells and high-grade lymphomas. In the present study, a series of Burkitt's lymphoma (BL) cell lines were analyzed for their expression of TIMP-1. TIMP-1 expression correlates with upregulation of activation and survival markers. TIMP-1-negative cells express the phenotype associated with group I BL lines and Epstein-Barr virus (EBV)-negative, nonendemic BLs (CD10+, CD38+, sIg+, and CD77+). However, TIMP-1+ BL lines showed group II/III BL phenotype, downregulation of the above markers, and upregulation and secretion of the activation marker CD23. Also, TIMP-1+ cells have high levels of CD40 expression. To determine whether TIMP-1 is directly involved in the BL phenotype, an EBV-negative BL line JD38 was infected with timp-1-expressing retrovirus and analyzed. In the absence of EBV, upregulation of TIMP-1 is sufficient to induce the same phenotype seen in TIMP-1+, EBV+ BL lines (CD10-, CD38-, sIg-, CD77-, CD23+, CD40 bright). This study not only suggests a role for TIMP-1 in BLs, but also supports its value as a prognostic factor. This is a US government work. There are no restrictions on its use.     

Role of T cells and germinal center formation in the generation of immune responses to the thymus-independent carbohydrate dextran B512

Immunization with the thymus-independent (TI) Ag native dextran (DX) B512 induces germinal center (GC) formation in the spleen. However, despite this GC formation, the anti-DX response is poor, and no affinity maturation can be observed. Using cholera toxin (CT) as an adjuvant, splenic as well as humoral responses to DX are improved. In this study, we investigated immune responses against DX in mice lacking TNF receptor I and in athymic mice. The adjuvant effect of CT on these responses was also evaluated. Mice lacking the TNF receptor I allowed us to investigate the role of follicular dendritic cell networks and GC formation in the spleen for the generation of Ab responses to DX, whereas we could investigate the role of T cells in GC development to TI Ags using athymic mice. We found that the humoral immune response to TI DX B512 was not dependent upon T cells or the presence of GCs, although GC development occurred after DX immunization. However, T cells were required for this GC formation, since athymic mice could not develop GCs after immunization with DX. We also show that even if CT is able to directly activate B cells when administered as an adjuvant, the major effect may require T cell participation; this is also the case for TI Ags. In contrast, CT adjuvancy is independent of GC formation.     

Expression level of a transgenic lambda2 chain results in isotype exclusion and commitment to B1 cells

Two new lambda2 chain-transgenic mouse lines were established, both of which showed stable transgene expression during aging of the mice. The line L23, which expressed the transgene at low levels, exhibited normal B cell development, antibody responses and serum Ig levels. Most of the B cells in this mouse line co-expressed the transgenic lambda2 chain together with an endogenous kappa chain, thus showing poor allelic exclusion of endogenous L chains. On the other hand, high expression of the transgenic lambda2 chain in the other mouse line, L2, resulted in nearly complete exclusion of endogenous L chain isotypes. In this line, the lambda2 transgene was already detectable in the cytoplasm of all preB-II cells and some pro/preB-I cells. Its expression during these early phases obviously inhibited development of conventional B2 cells, since the B cells in the periphery of these mice were almost exclusively of the B1 type. This finding was confirmed by adoptive transfer of transgenic bone marrow into lethally irradiated recipients. Very few B cells were present in the spleen of such recipients. The serum IgM levels of L2 mice were close to normal and the majority of these IgM were associated with the transgenic lambda2 chain. Antibody responses to thymus-dependent antigens in such mice were almost exclusively found to be of IgM class. Together, these findings indicate a developmental bias leading to a predominance of B1 cells in the L2 line.     

Distinct UV-B and UV-A/blue light signal transduction pathways induce chalcone synthase gene expression in Arabidopsis cells

UV and blue light control the expression of flavonoid biosynthesis genes in a range of higher plants. To investigate the signal transduction processes involved in the induction of chalcone synthase (CHS) gene expression by UV-B and UV-A/blue light, we examined the effects of specific agonists and inhibitors of known signaling components in mammalian systems in a photomixotrophic Arabidopsis cell suspension culture. CHS expression is induced specifically by these wavelengths in the cell culture, in a manner similar to that in mature Arabidopsis leaf tissue. Both the UV-B and UV-A/blue phototransduction processes involve calcium, although the elevation of cytosolic calcium is insufficient on its own to stimulate CHS expression. The UV-A/blue light induction of CHS expression does not appear to involve calmodulin, whereas the UV-B response does; this difference indicates that the signal transduction pathways are, at least in part, distinct. We provide evidence that both pathways involve reversible protein phosphorylation and require protein synthesis. The UV-B and UV-A/blue light signaling pathways are therefore different from the phytochrome signal transduction pathway regulating CHS expression in other species.     

The effect of angiotensin II on myosin heavy chain expression in cultured myocardial cells

Angiotensin II (AII), the principal mediator of the renin-angiotensin system, is an important regulator of vascular and cardiac homeostasis. AII has also been shown to be a regulator of cardiac hypertrophy and of the corresponding changes in amount and composition of certain tissue proteins. We examined the trophic effects of AII on cultured myocytes derived from neonatal rat ventricles and followed, by Northern blot analysis and polyacrylamide gel electrophoresis, the expression of alpha- and beta-myosin heavy chain iso-mRNAs and isoproteins. Our findings show that a single administration of AII is sufficient to induce a trophic response in cultured beating myocytes and to enhance the expression of beta-myosin heavy chain iso-mRNA and isoprotein, having no effect on alpha-myosin heavy chain. Induction of alpha-myosin heavy chain expression by thyroid hormone before AII was administered showed that AII could not potentiate a shift from alpha- to beta-myosin heavy chain predominance. We suggest that the potency of AII to regulate the expression of myosin heavy chain isogenes is restricted to the beta isoform and is overridden by thyroid hormone.     

The BAX gene, the promoter of apoptosis, is mutated in genetically unstable cancers of the colorectum, stomach, and endometrium

OBJECTIVE: The study's objective was to determine whether there is a difference in the plasma concentration of adrenomedullin, a hypotensive peptide, between arterial and venous umbilical cord blood of uncomplicated gestations with vaginal delivery. STUDY DESIGN: Arterial and venous umbilical cord blood was obtained immediately after vaginal delivery of 44 term infants with uncomplicated antepartum and intrapartum courses. Radioimmunoassay was performed to assess adrenomedullin concentrations in the plasma. The paired t test was used to compare arterial and venous concentrations. Significance was set at P < .05. RESULTS: Mean +/- SE adrenomedullin concentrations were 178.7 +/- 4.7 pg/mL and 190.6 +/- 6.3 pg/mL for arterial and venous cord plasma, respectively. The difference between the 2 concentrations was not significant (11.8 pg/mL, P = .09). CONCLUSION: Arterial and venous umbilical plasma concentrations of adrenomedullin do not differ significantly in uncomplicated gestations terminating with uncomplicated vaginal deliveries. This suggests that in the normal state there is neither net production nor net clearance of adrenomedullin in the placenta.