δ-Cadinene inhibits the growth of ovarian cancer cells via caspase-dependent apoptosis and cell cycle arrest

Ovarian cancer is one of the most common causes of mortality among all cancers in females and is the primary cause of mortality from gynecological malignancies. The objective of the current research work was to evaluate a naturally occurring sesquiterpene-δ-Cadinene for its antiproliferative and apoptotic effects on human ovary cancer (OVCAR-3) cells. We also demonstrated the effect of δ-Cadinene on cell cycle phase distribution, intracellular damage and caspase activation. Sulforhodamine B (SRB) assay was used to evaluate the antiproliferative effect of δ-cadinene on OVCAR-3 cells. Cellular morphology after δ-cadinene treatment was demonstrated by inverted phase contrast microscopy, fluorescence microscopy and transmission electron microscopy. Flow cytometry was used to analyze the effect of δ-cadinene on cell cycle phase distribution and apoptosis using propidium iodide and Annexin V-fluorescein isothiocyanate (FITC)/PI kit. The results revealed that δ-cadinene induced dose-dependent as well as time-dependent growth inhibitory effects on OVACR-3 cell line. δ-cadinene also induced cell shrinkage, chromatin condensation and nuclear membrane rupture which are characteristic of apoptosis. Treatment with different doses of δ-cadinene also led to cell cycle arrest in sub-G1 phase which showed dose-dependence. Western blotting assay revealed that δ-cadinene led to activation of caspases in OVCAR-3 cancer cells. PARP cleavage was noticed at 50 µM dose of δ-cadinene with the advent of the cleaved 85-kDa fragment after exposure to δ-cadinene. At 100 µM, only the cleaved form of PARP was detectable. Pro-caspase-8 expression remained unaltered until 10 µM dose of δ-cadinene. However, at 50 and 100 µM dose, pro-caspase-8 expression was no longer detectable. There was a significant increase in the caspase-9 expression levels after 50 and 100 µM δ-cadinene treatments.     

Flow Cytometric Determination of Ganciclovir Susceptibilities of Human Cytomegalovirus Clinical Isolates

A flow cytometric assay has been developed for the measurement of susceptibilities to ganciclovir of laboratory strains and clinical isolates of human cytomegalovirus (HCMV). The assay uses fluorochrome-labeled monoclonal antibodies to HCMV immediate-early and late antigens to identify HCMV-infected cells and flow cytometry to detect and quantitate the number of antigen-positive cells. By this assay, the 50 and 90% inhibitory concentrations (IC₅₀ and IC₉₀, respectively) of ganciclovir for the AD169 strain of HCMV were 1.7 and 9.2 μM, respectively, and the IC₅₀ for the ganciclovir-resistant D6/3/1 derivative of the AD169 strain was greater than 12 μM. The ganciclovir susceptibilities of 17 HCMV clinical isolates were also determined by flow cytometric analysis of the effect of ganciclovir on late-antigen synthesis in HCMV-infected cells. The average IC₅₀ of ganciclovir for drug-sensitive HCMV clinical isolates was 3.79 μM (±2.60). The plaque-reduction assay for these clinical isolates yielded an average IC₅₀ of 2.80 μM (±1.46). Comparison of the results of the flow cytometry assays with those obtained from the plaque-reduction assays demonstrated acceptable bias and precision. Flow cytometric and plaque-reduction analysis of cells infected with ganciclovir-resistant clinical isolates failed to show a reduction in the percentage of late-antigen-positive cells or PFU, even at 96 μM ganciclovir. The flow cytometric assay for determining ganciclovir susceptibility of HCMV is quantitative, and objective, and potentially automatable, and its results are reproducible among laboratories.     

Dihydromethysticin kavalactone induces apoptosis in osteosarcoma cells through modulation of PI3K/Akt pathway,disruption of mitochondrial membrane potential and inducing cell cycle arrest

The objective of the present study was to evaluate the tumor and apoptotic effects of dihydromethysticin kavalactone against human osteosarcoma (MG-63) cells. Antiproliferative activity was measured with the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Apoptosis induction by dihydromethysticin was demonstrated by fluorescence microscopy, quantitative videomicroscopy and Annexin V-FITC apoptosis detection kit. Mitochondrial membrane potential disruption was demonstrated by rhodamine-123 dye using flow cytometry. We also evaluated the effect of dihydromethysticin on PI3K/Akt pathway with an immunoblotting analysis. The results showed that the compound induced dose-dependent as well as time-dependent antiproliferative effects against MG-63 cell growth. Cell death and apoptotic body formation was noticed followed dihydromethysticin treatment at various doses. The percentage of apoptotic cells (early apoptosis + late apoptosis) increased from 6.63% in untreated control to 23.92%, 23.81% and 93.9% in 25 µM, 75 µM and 100 µ Mdihydromethysticin-treated cells respectively. Flow cytometric analysis showed dihydromethysticin induced an increase in G0/G1 cells (apoptotic cells). Furthermore, we observed mitochondrial transmembrane depolarization along with decreased phosphorylation levels for PI3K, AKT (Ser 473), AKT (Thr 308), GSK-3β, and BAD. These reductions were associated with down regulation of AKT and upregulation of both GSK-3β and BAD.     

High α-defensin levels in patients with systemic lupus erythematosus

Innate immunity plays a role in systemic lupus erythematosus (SLE). Our objective was to determine the levels of defensins, which are antimicrobial and immunomodulatory polypeptides, in SLE. Sera from SLE patients and healthy controls were tested for pro-inflammatory human β-defensin 2 (hBD-2) and for α-defensin human neutrophil peptide 1 (HNP-1). hBD-2 could not be detected by enzyme-linked immunosorbent assay (ELISA) and its mRNA levels were low in SLE patients and similar to those found in controls. In contrast, the mean α-defensin level in the sera of all SLE patients (11·07 ± 13·92 ng/μl) was significantly higher than that of controls (0·12 ± 0·07 ng/μl). Moreover, 60% of patients demonstrated very high serum levels (18·5 ± 13·36 ng/μl) and 50% showed elevated gene expression in polymorphonuclear cells. High α-defensin levels correlated with disease activity, but not with neutrophil count. Thus, activation and degranulation of neutrophils led to α-defensin secretion in SLE patients. Given the immunomodulatory role of α-defensins, it is possible that their secretion may activate the adaptive immune system leading to a systemic response.     

γ-terpineol inhibits cell growth and induces apoptosis in human liver cancer BEL-7402 cells in vitro

To investigate the effect of γ-terpineol on cell proliferation and apoptosis of human hepatoma BEL-7402 cells to elucidate its molecular mechanism. Here, BEL-7402 cells were treated with various concentrations (40, 80, 160, 320 and 640 μg/ml) of γ-terpineol for 48 h, cell proliferation was determined by 3-(4,5-dimethyl-thiazolyl-2)-2,5-diphenyl tetrazolium bromides (MTT) assay. Cell colony inhibition was determined by soft agar assay. Apoptosis and possible molecular mechanisms were evaluated by morphological observation, flow cytometry analysis, and DNA fragmentation assay. The γ-terpineol significantly suppressed BEL-7402 cell proliferation in a dose-dependent manner. Characteristic morphological and biochemical changes associated with apoptosis such as cells shrinkage, deformation and vacuolization of mitochondria, nuclear chromatin condensation and fragmentation, formation of apoptotic bodies were observed after BEL-7402 cells treated with γ-terpineol for 24 h and 48 h. Cell cycle were displayed by flow cytometry analysis, the γ-terpineol treatment resulted in accumulation of cells at G₁ or S phase and a blockade of cell proliferation compared to control group. Treating BEL-7402 cells with 320 μg/ml of γ-terpineol for 36 h and 48 h, a typical apoptotic “DNA ladder” was observed using DNA fragmentation assay. The present study demonstrated that possible anti-cancer mechanism of γ-terpineol on human hematomas cells is through inducing cell apoptosis to suppress tumor cell growth.     

Effect of rTsP53 on the M1/M2 activation of bone-marrow derived macrophage in vitro

We investigated that if rTsP53 could be used to activate bone-marrow derived macrophage (BMDM) into M2 macrophage and stop M1 macrophage activation. After 72 h incubation in blank culture medium, cells with PE-CCR7 (-) and FITC-CD206 (-) was extracted and its mean proportion was 92.30 ± 0.22%. With the stimulation of 20 μg/ml IFN-γ for 72 h, cells with PE-CCR7 (+) was extracted and its mean proportion was 16.24 ± 0.82%. With the stimulation of IL-3/IL-14 (both 10 μg/ml) for 72 h, cells with FICT-CD206 (+) was extracted and its mean proportion was 87.32 ± 4.29%. Co-incubation with different dose of rTsP53 (0.001 μg/ml, 0.01 μg/ml, 0.1 μg/ml, 1 μg/ml, 2 μg/ml, 5 μg/ml, 10 μg/ml, respectively) for 72 h, FITC-CD206 (+) macrophage was extracted. The mean proportion in each group was 1.09 ± 0.22%, 2.13 ± 0.13%, 4.91 ± 0.07%, 5.48 ± 0.29%, 9.81 ± 0.06%, 12.83 ± 0.55%, 17.87 ± 0.02%, respectively. The dose of rTsP53 was significantly positive correlated to the proportion of FITC-CD206 (+) macrophage. Co-incubation with 20 μg/ml IFN-γ and 5 μg/ml rTsP53 for 72 h, cells with PE-CCR7 (+) was extracted and its mean proportion was 10.60 ± 0.19%. Compared to that of mere co-incubation with IFN-γ, there was significant difference between the two groups. ELISA showed that Th1 cytokines’ (IFN-γ, IL-6 and TNF-α) level decreased in the culture medium supernatant of BMDM co-incubated with rTsP53. There was negative correlation between the Th1 cytokines’ level and the dose of rTsP53. Both Th2 cytokines (IL-4 and IL-13) and regulatory cytokines in the culture medium increased. There was positive correlation between the Th2 cytokines’ level and the dose of rTsP53. There was also positive correlation between the regulatory cytokines’ level and the dose of rTsP53. Compared to that of BMDM co-incubated with IFN-γ, levels of TNF-α and IL-6 were significant lower than that of BMDM co-incubated with both IFN-γ and rTsP53 (both P < 0.05), while the levels of IL-4 and TGF-β were significant higher (both P < 0.05). There was no significant difference in the levels of IL-13 and IL-10 between the two groups.     

In vitro inhibitory effects of imatinib mesylate on stromal cells and hematopoietic progenitors from bone marrow

Imatinib mesylate (IM) is used to treat chronic myeloid leukemia (CML) because it selectively inhibits tyrosine kinase, which is a hallmark of CML oncogenesis. Recent studies have shown that IM inhibits the growth of several non-malignant hematopoietic and fibroblast cells from bone marrow (BM). The aim of the present study was to evaluate the effects of IM on stromal and hematopoietic progenitor cells, specifically in the colony-forming units of granulocyte/macrophage (CFU-GM), using BM cultures from 108 1.5- to 2-month-old healthy Swiss mice. The results showed that low concentrations of IM (1.25 µM) reduced the growth of CFU-GM in clonogenic assays. In culture assays with stromal cells, fibroblast proliferation and α-SMA expression by immunocytochemistry analysis were also reduced in a concentration-dependent manner, with a survival rate of approximately 50% with a dose of 2.5 µM. Cell viability and morphology were analyzed using MTT and staining with acrydine orange/ethidium bromide. Most cells were found to be viable after treatment with 5 µM IM, although there was gradual growth inhibition of fibroblastic cells while the number of round cells (macrophage-like cells) increased. At higher concentrations (15 µM), the majority of cells were apoptotic and cell growth ceased completely. Oil red staining revealed the presence of adipocytes only in untreated cells (control). Cell cycle analysis of stromal cells by flow cytometry showed a blockade at the G₀/G₁ phases in groups treated with 5-15 µM. These results suggest that IM differentially inhibits the survival of different types of BM cells since toxic effects were achieved.     

Rhein induces apoptosis of human gastric cancer SGC-7901 cells via an intrinsic mitochondrial pathway

Rhein is a primary anthraquinone found in the roots of a traditional Chinese herb, rhubarb, and has been shown to have some anticancer effects. The aim of the present study was to investigate the effect of rhein on the apoptosis of the human gastric cancer line SGC-7901 and to identify the mechanism involved. SGC-7901 cells were cultured and treated with rhein (0, 50, 100, 150, and 200 µM) for 24, 48, or 72 h. Relative cell viability assessed by the MTT assay after treatment was 100, 99, 85, 79, 63% for 24 h; 100, 98, 80, 51, 37% for 48 h, and 100, 97, 60, 36, 15% for 72 h, respectively. Cell apoptosis was detected with TUNEL staining and quantified with flow cytometry using annexin FITC-PI staining at 48 h after 100, 200 and 300 µm rhein. The percentage of apoptotic cells was 7.3, 21.9, 43.5%, respectively. We also measured the mRNA levels of caspase-3 and -9 using real-time PCR. Treatment with 100 µM rhein for 48 h significantly increased mRNA expression of caspase-3 and -9. The levels of apoptosis-related proteins including Bcl-2, Bax, Bcl-xL, and pro-caspase-3 were evaluated in rhein-treated cells. Rhein increased the Bax:Bcl-2 ratio but decreased the protein levels of Bcl-xL and pro-caspase-3. Moreover, rhein significantly increased the expression of cytochrome c and apoptotic protease activating factor 1, two critical components involved in mitochondrial pathway-mediated apoptosis. We conclude that rhein inhibits SGC-7901 proliferation by inducing apoptosis and this antitumor effect of rhein is mediated in part by an intrinsic mitochondrial pathway.     

Kaurene diterpene induces apoptosis in U87 human malignant glioblastoma cells by suppression of anti-apoptotic signals and activation of cysteine proteases

Gliomas are the most common and malignant primary brain tumors in humans. Studies have shown that classes of kaurene diterpene have anti-tumor activity related to their ability to induce apoptosis. We investigated the response of the human glioblastoma cell line U87 to treatment with ent-kaur-16-en-19-oic acid (kaurenoic acid, KA). We analyzed cell survival and the induction of apoptosis using flow cytometry and annexin V staining. Additionally, the expression of anti-apoptotic (c-FLIP and miR-21) and apoptotic (Fas, caspase-3 and caspase-8) genes was analyzed by relative quantification (real-time PCR) of mRNA levels in U87 cells that were either untreated or treated with KA (30, 50, or 70 µM) for 24, 48, and 72 h. U87 cells treated with KA demonstrated reduced viability, and an increase in annexin V- and annexin V/PI-positive cells was observed. The percentage of apoptotic cells was 9% for control cells, 26% for cells submitted to 48 h of treatment with 50 µM KA, and 31% for cells submitted to 48 h of treatment with 70 µM KA. Similarly, in U87 cells treated with KA for 48 h, we observed an increase in the expression of apoptotic genes (caspase-8, -3) and a decrease in the expression of anti-apoptotic genes (miR-21 and c-FLIP). KA possesses several interesting properties and induces apoptosis through a unique mechanism. Further experiments will be necessary to determine if KA may be used as a lead compound for the development of new chemotherapeutic drugs for the treatment of primary brain tumors.     

Human peripheral blood T lymphocytes express α-, μ- and F(ab')2-related determinants in their surface membranes

Whereas a mean of 81% of freshly isolated human T cells bound purified chicken anti-F(ab')₂ antibodies to their surface membranes, 14 and 2% bound chicken anti-μ and chicken anti-α antibody preparations respectively as demonstrated by the mixed antiglobulin rosetting reaction (MARR). Neuraminidase treatment of T cells significantly increased their reactivity in the MARR so that an average of 98, 39 and 40% bound anti-F(ab')₂ anti-μ and anti-α reagents respectively. When using anti-F(ab')₂ antibodies, inclusion of Fabγ in the rosetting medium caused greater than 90% inhibition of the MARR whereas Fcγ produced only 25% inhibition; this indicates that the determinants seen by anti-F(ab')₂ antibodies are not carbohydrate in nature since Fabγ and Fcγ fragments prepared from a human IgG myeloma express identical carbohydrate moieties. To discover whether the various immunoglobulin (Ig) related T cell molecules play a role in antigen recognition, each antiglobulin reagent was assessed for its capacity to inhibit T cell binding of a selected antigen, i.e. fluorescein-labelled keyhole limpet haemocyanin. Whereas the frequency of antigen-binding lymphocytes (ABL) varied from 8·1 to 15·1 per 10³ cells, fewer than 1 in 10³ cells were Ig-positive with a rabbit F(ab')₂ anti-light chain reagent. Thus virtually all ABL were T cells. Each antiglobulin reagent produced 50% or greater inhibition of antigen binding thereby suggesting that F(ab')₂-, α-, and μ-related determinants are in close proximity to, or part of, the T cell antigen receptor. More substantial evidence for a direct association of Ig-related surface determinants and the T cell antigen receptor is provided by the finding that under capping conditions, modulation of surface F(ab')₂-related determinants by anti-F(ab')₂ antibodies reduced antigen binding. Further, that anti-F(ab')₂ antibodies induced F(ab')₂-, α- and μ-related determinants to co-cap suggests that F(ab')₂-related determinants are stably associated with α- and/or μ-related determinants in the T cell membrane and are part of the same Ig-related molecule(s).     

Inhibition of ovarian cancer proliferation and invasion by pachymic acid

To determine the effect of pachymic acid (PA) on proliferation, cell cycle, and invasion in human ovarian carcinoma cell lines HO-8910 and explore some possible mechanisms, HO-8910 cells was treated with different concentrations of PA (0.5, 1, 2 μM). CCK-8 assay, propidium iodide staining, was applied to measuring the growth inhibiting rates of HO-8910 cells. Cell cycle was measured by flow cytometry. In addition, the activity of PA against HO-8910 cells invasion was evaluated in transwell assay. Western blot detected the proteins expression of E-cadherin, β-catenin and COX-2 of different groups treated with PA in different concentrations (0.5, 1, 2 μM) for 48 h. Our results showed that PA could effectively inhibit the in vitro growth of HO-8910 cells in dose-dependent manners in 72 h, suppressed migration and invasion of HO-8910 cells in concentration-dependent manners at 24 h, caused the increased accumulation of G1 phase cells, and caused down-regulation of β-catenin and COX-2 and up-regulation of E-cadherin expression level. Taken together, it could conclude that PA might inhibit proliferation and invasion of ovarian carcinoma cell through decreasing β-catenin and COX-2 expression and increasing E-cadherin exprssion.     

Effects of portulacerebroside a on apoptosis of human leukemia HL60 cells and p38/JNK signaling pathway

Acute myeloid leukemia is known as one of the most malignant diseases. We aimed at exploring the effect of portulacerebroside A (PCA) on the apoptosis in human leukemia HL60 cells and clarify the possible mechanisms involved in. By MTT analysis, we found that PCA (1-100 μM) inhibited the cell viability in a time- and dose-dependent manner, and cell cycle was arrested at G0/G1 period. PCA treatment from 5 to 50 μM dose-dependently induced apoptosis from 12.7 ± 1.56% to 52.7 ± 6.214% of HL60 cells. Mitochondrial membrane potential (MMP) was decreased and reactive oxygen species (ROS) accumulated obviously. mRNA expressions and protein levels of Bax/Bcl-2, caspase-3 and caspase-9 were elevated significantly. ERK1/2, JNK1/2 and p38 MAPK pathway were blocked detected by western blot analysis. In conclusion, PCA can act as a new agent for leucocythemia treatment.     

Relevance of 123I-BMIPP delayed scintigraphic imaging for patients with angina pectoris – a pilot study

Introduction

The study was designed to clarify the role of ¹²³I-β-methyl-iodophenylpentadecanoic acid (¹²³I-BMIPP) in the evaluation of myocardial fatty acid metabolism in patients with stable angina pectoris (AP) before and after percutaneous coronary intervention (PCI).

Material and methods

Ten controls (mean age: 70.4 ±10.5 years) and 12 patients with AP (mean age: 67.4 ±11.6 years) and single vessel coronary artery disease participated in the radionuclide cardiac study. Scintigraphic images were acquired at 30 min and at 4 h after ¹²³I-BMIPP injection to determine early and delayed BMIPP uptake, respectively. The heart-to-mediastinum (H/M) ratio and the washout rate (WR) were calculated from the planar images. All patients underwent scintigraphy one day before PCI and again 1 month after successful PCI.

Results

No significant differences in the early or delayed H/M ratios were observed between the patients and the controls before PCI (early: 2.70 ±0.36 vs. 2.73 ±0.57; delayed: 2.26 ±0.33 vs. 2.40 ±0.43; p > 0.2 for both). The early and delayed H/M ratios remained unchanged with the comparison with before PCI (early: 2.72 ±0.27, delayed: 2.23 ±0.22; p > 0.2 for both). The global WR before PCI was significantly higher in the patients than in the control group (36.7 ±9.3%, vs. 28.1 ±8.2%, p = 0.02). However, the WR after PCI did not significantly differ between the patients and the controls (34.3 ±7.8% vs. 28.1 ±8.2%, p = 0.1).

Conclusions

These data may suggest that the WR of ¹²³I-BMIPP determined from the planar images enhances the presence of myocardial ischaemia.     

Radiosensitivity of human ovarian carcinoma and melanoma cells to γ-rays and protons

Introduction

Proton radiation offers physical advantages over conventional radiation. Radiosensitivity of human 59M ovarian cancer and HTB140 melanoma cells was investigated after exposure to γ-rays and protons.

Material and methods

Irradiations were performed in the middle of a 62 MeV therapeutic proton spread out Bragg peak with doses ranging from 2 to 16 Gy. The mean energy of protons was 34.88 ±2.15 MeV, corresponding to the linear energy transfer of 4.7 ±0.2 keV/µm. Irradiations with γ-rays were performed using the same doses. Viability, proliferation and survival were assessed 7 days after both types of irradiation while analyses of cell cycle and apoptosis were performed 48 h after irradiation.

Results

Results showed that γ-rays and protons reduced the number of viable cells for both cell lines, with stronger inactivation achieved after irradiation with protons. Surviving fractions for 59M were 0.91 ±0.01 for γ-rays and 0.81 ±0.01 for protons, while those for HTB140 cells were 0.93 ±0.01 for γ-rays and 0.86 ±0.01 for protons. Relative biological effectiveness of protons, being 2.47 ±0.22 for 59M and 2.08 ±0.36 for HTB140, indicated that protons provoked better cell elimination than γ-rays. After proton irradiation proliferation capacity of the two cell lines was slightly higher as compared to γ-rays. Proliferation was higher for 59M than for HTB140 cells after both types of irradiation. Induction of apoptosis and G2 arrest detected after proton irradiation were more prominent in 59M cells.

Conclusions

The obtained results suggest that protons exert better antitumour effects on ovarian carcinoma and melanoma cells than γ-rays. The dissimilar response of these cells to radiation is related to their different features.     

Increased Th2-like Invariant Natural Killer T cells in Peripheral Blood From Patients With Asthma

Purpose

Invariant natural killer T (iNKT) cells might play an important role in asthma pathogenesis in humans. Our previous study found no difference in the number of blood iNKT cells between asthma patients and controls. However, few studies have examined the function of blood iNKT cells in human asthma.

Methods

Twenty asthma patients and eight controls were included in this study. Blood iNKT cells were identified using double staining with anti-Vα24 and anti-Vβ11 monoclonal antibodies (mAbs) or with 6B11 and anti-Vβ11 mAbs. Intracellular IL-4, IL-10, and IFN-γ cytokines were stained in blood iNKT cells using their respective mAbs and isotypes. In addition, their relationships with clinical parameters were analyzed.

Results

The number of Vα24+Vβ11+ iNKT cells or 6B11+Vβ11+ iNKT cells did not differ between asthma patients and controls. However, among Vα24+Vβ11+iNKT cells, the proportion of IL-4+iNKT cells was increased in asthma patients compared to controls (7.0±3.0% vs 0.5%±0.4%, P<0.05). There were no differences in the proportions of IL-10+or IFN-γ+iNKT cells between the groups. The proportion of IL-4+ cells among 6B11+Vβ11+iNKT cells inversely correlated with FEV1, expressed as a percentage predicted value in asthma patients (Rs=-0.64, P<0.05, n=19).

Conclusions

Blood iNKT cells are thought to be Th2-like, and IL-4-producing iNKT cells may be associated with lung function in human asthma.     

Membrane-associated immunoglobulins of human lymphocytes in immunologic disorders

Membrane-associated immunoglobulins of peripheral blood lymphocytes were studied by indirect immunofluorescence for γ, α, μ, κ and λ chains in healthy subjects and patients with immunologic disease.

In healthy subjects, heavy chains were found on 30·7% of lymphocytes (γ 15·3%, α 7·2% and μ 8·2%) and light chains on 32·8% of cells (κ 20·4% and λ 12·4%). Patients with humoral immune deficiencies had fewer immunoglobulin-bearing cells; sarcoidosis or thymectomy patients had normal or decreased immunoglobulin-bearing lymphocytes; cells with light chains were fewer than those with heavy chains on their lymphocytes. In some cases, normal levels of serum immunoglobulins were found in the absence of the corresponding immunoglobulin-bearing cells, and in others normal immunoglobulin-bearing lymphocytes were present in the absence of the corresponding serum immunoglobulins.

These data suggest that (1) immunoglobulin-bearing lymphocytes in blood do not reflect the condition of immunoglobulin-synthesizing cells in peripheral lymphoid tissues, and (2) in certain immunologic disorders, either some B-lymphocytes do not synthesize immunoglobulins, or immunoglobulins are in such a situation that the whole molecule or part of the molecule is not visualized by current methods.

     

Increased numbers of CD5+ B cells and T cell receptor (TCR) γδ+ T cells are associated with younger age of onset in rheumatoid arthritis (RA)

Patients presenting with RA before the age of 45 years (younger onset) are known to have more aggressive disease compared with patients presenting after the age of 65 years (older onset). Coordinated expansion of circulating CD5⁺ B cell and TCR γδ⁺ T cell levels has been reported in patients with RA. This study assesses the peripheral blood levels of these two cell types in RA patients with younger and older onset of disease. CD5⁺ B cell levels were significantly elevated in the younger onset RA group (26·6 ± 4·5%) compared with the older onset RA group (14·2 ± 1·2%; P <0·01). TCR γδ⁺ T cell levels were also significantly raised in the young patients (4·0 ± 0·9%) compared with elderly patients (1·6 ± 0·2%; P <0·01). T cell levels (CD3⁺) were similar in both groups (young 66·4 ± 3·3%; old 74·3 ± 3·4% (mean ± s.e.m.); NS). Total B cell levels (CD19⁺) were also similar in these groups (7·7 ± 0·7% versus 8·9 ± 1·8%; NS). A significant positive correlation was observed between the CD5⁻ B and TCR γδ⁺ T cell types in the patients (r = 0·72, P <0.05). Compared with age-matched normal controls, the younger onset patients had similar CD5⁺ B cell and TCR γδ⁺ T cell levels to the elderly controls (CD5⁺ B cells 30·2 ± 3·0%; TCR γδ⁺ T cells 3·0 ± 0·8%). Conversely, older onset RA patients had CD5⁺ B cell levels similar to the young controls (12·3 ± 1·9%). Spontaneous in vitro synthesis of immunoglobulins (IgM, IgA and IgG) and rheumatoid factors (IgM and IgA isotypes) were not significantly different in both patient groups. The coordinate expansion of circulating CD5⁺ B cells and γδ⁺ T cells seen in patients with RA presenting before 45 years of age and not after 65 years of age may suggest a potential role for these cells in more aggressive disease states.     

Kappa and lambda receptor sites on single lymphocytes

Light chain receptors on human circulating lymphocytes of thirty-three normal individuals were visualized by the immunocyto-adhesion reaction in which human (anuclear) erythrocytes coated with κ chains and chicken (nucleated) erythrocytes coated with λ chains (or vice versa) were used. 4·4 (±2·3)% of the lymphocytes had κ and 4·15 (±2·4)% had λ receptors, when the lymphocytes were sensitized with antisera to either κ and λ chains. When both antisera were used simultaneously only 5·3 (±3)% were found to have receptors and the majority of cells carried both κ and λ receptors. These findings suggest that in these cells the genes for the constant regions of both light chains are active.