Basic types and vital forms of human peripheral and PHA-stimulated lymphocytes studied in vitro

Natural diversity in peripheral and PHA-stimulated lymphocytes seen in the same donors was studied using digitized streak photo of living cells in observational camera. Cells were monitored for 5-8 h at the superior limit of optical resolution by means of phase-contrast microscopy. Intact lymphocytes were observed in autological blood plasma, and PHA-stimulated lymphocytes were examined in self-conditioned centrifuged growth medium. The majority of intact cells were small- and middle-sized floating lymphocytes with microvilli, and middle-sized caudate lymphocytes capable of stick-slip motion. The lesser part consisted of "spread-eagle" or movable forms of both large granular lymphocytes and middle-sized lymphocytes of several types: narrow-plasm lymphocytes with lamellipodia, wide-plasm lymphocytes without cytoplasmic processes, lymphocytes with single pseudopodia, and lymphocytes with single lobopodia of complex shape. On the contrary, the minor fraction of PHA-stimulated lymphocytes of 3 day old cultures contained floating cells with microvilli or floating cells with microvilli and two pseudopodia, whereas the majority of these lymphocytes were spread-eagle or movable forms of cells of different type. These substrate-adhesive PHA-stimulated lymphocytes had well defined apical and basal cell surfaces, but upon mechanical stress are easily pinched off to become ball-shaped. At least 6 different cell types were distinguished among substrate-adhesive PHA-stimulated lymphocytes, with more than half of these being heavily vacuolated spheroid lymphocytes prone to forming cell clusters. The rest PHA-stimulated lymphocytes were represented by signet-ring lymphocytes with dark or light cytoplasm, narrow-plasm lymphocytes with large prolonged nuclei and lamellipodia, lymphocytes with single lobopodia, and lymphocytes with single spiral structures in the cytoplasm. The spiral structure is 10-11 microns in length and 0.5-0.7 micron in width, being presumably a mitochondrion or a group of butt-joined mitochondria. Since some of the caudate middle-sized lymphocytes also contain this structure, these may be regarded as putative precursors of respective type of PHA-stimulated lymphocytes. Under the conditions of observation, interphase nuclei of all live PHA-stimulated lymphocytes were seen to contain numerous globular or fiber structures of condensed chromatin made of 0.3-0.8 micron beads. These beads are doubtless interphase chromomeres.     

Concentration of nucleotides and deoxynucleotides in peripheral and phytohemagglutinin-stimulated mammalian lymphocytes. Effects of adenosine and deoxyadenosine

Concentrations of purine and pyrimidine ribonucleotides were measured with HPLC in lymphocytes of man, horse, pig and sheep and in rat thymocytes. The ATP concentration was highest in lymphocytes of all species and about 850 pmol/10(6) cells in human and equine lymphocytes, higher in porcine and lower in ovine lymphocytes and rat thymocytes. The GTP concentration was comparable in human, equine and porcine lymphocytes, but lower in ovine lymphocytes. ATP concentration was also measured in lymphocytes of man, horse and pig with a luciferin-luciferase assay. During culturing with or without phytohemagglutinin the ATP concentrations decreased in these lymphocytes. The concentrations of TTP and dATP were measured with a DNA polymerase assay. Phytohemagglutinin-stimulation increased the TTP concentration in lymphocytes of all three species, the dATP concentration only in human lymphocytes. ATP, TTP and dATP concentrations and thymidine incorporation were measured in phytohemagglutinin-stimulated lymphocytes after 24 and 48 h culturing in the presence of adenosine or deoxyadenosine. Adenosine increased the ATP concentration in porcine and equine, but not in human lymphocytes. Deoxyadenosine and adenosine did not affect the TTP concentration. Deoxyadenosine decreased the ATP concentration only in the presence of EHNA in human lymphocytes, but increased it in other conditions and in equine and porcine lymphocytes. Deoxyadenosine in the presence of EHNA increased the dATP concentration in human, equine and porcine lymphocytes 3-, 10-, and 9-fold, respectively, and decreased considerably thymidine incorporation. Deoxyadenosine without EHNA increased the dATP concentration 2-5-fold, decreased the thymidine incorporation in lymphocytes of man and horse, but stimulated incorporation in porcine lymphocytes about 5-fold. The latter results indicate that accumulation of dATP is not always associated with inhibition of cell proliferation.     

Increased T lymphocytes and IgMEA-receptor lymphocytes in Hodgkin's disease spleens.

Splenic lymphocytes from 11 patients with Hodgkin's disease were compared to lymphocytes of six spleens from patients with nonlymphoproliferative diseases. T lymphocytes were increased in patients with histological involvement by Hodgkin's disease. Likewise, lymphocytes from spleens with histological involvement showed increased rosette formation with immunologlobulin M-coated sheep red blood cells (IgMEA). A similar increase in T lymphocytes and in IgMEA rosette formation was not observed with normal peripheral blood lymphocytes, control spleens, or with Hodgkin's disease spleens without evidence of histological involvement.     

半滑舌鳎外周血T淋巴细胞的分离、鉴定及TCRβ基因免疫应答分析

为开展半滑舌鳎(Cynoglossus semilaevis)免疫学研究提供细胞平台, 利用密度梯度离心法分离半滑舌鳎外周血淋巴细胞, 采用短期细胞培养法分离悬浮淋巴细胞, 悬浮淋巴细胞在含有0.3 μg/mL的PHA的DMEM完全培养基, 于24℃条件下可连续培养3—4d左右, 采用自制的尼龙毛柱可将悬浮淋巴细胞中的非黏附淋巴细胞和黏附淋巴细胞成功分离; 利用流式细胞仪结合特异抗体检测对非黏附淋巴细胞和黏附淋巴细胞进行鉴定, 结果表明, 非黏附细胞与鼠抗人FTIC-CD3单克隆抗体特异结合, 为T样淋巴细胞; 黏附细胞和鼠抗人FTIC-CD19单抗特异结合, 为B样淋巴细胞。T细胞表面抗原受体TCRβ基因可特异性的在非黏膜细胞中表达, 而在黏附细胞中不表达, 证明分离获得的非黏膜细胞为T淋巴细胞, 采用qRT-PCR (Quantitative Real-Time PCR)方法检测TCRβ基因表达, 结果表明, TCRβ基因在半滑舌鳎肝、脾、头肾、后肾、小肠、胃、血液、鳃、皮肤、肌肉、心脏、脑、卵巢组织中均有表达, 其中在肠、胃、脾、头肾中表达量较高; 鳗弧菌感染后TCRβ基因在肝、脾、鳃中呈现明显的上调表达, 且表达峰值出现在感染后72—96h, 表明TCRβ基因在获得性免疫应答中起重要作用。     

Application of orthogonal light scattering for routine screening of lymphocyte samples

Orthogonal and forward light-scattering properties of lymphocytes were measured from patients with different lymphocytic diseases in order to determine the potential value of light scattering as a screening device. Monitoring of orthogonal light scattering of lymphocytes of a B-cell chronic lymphocytic leukemia patient during splenic irradiation (SI) revealed the selective decrease of malignant cells and the fact that the major part of the residual lymphocytes were cytotoxic lymphocytes. By combining forward and orthogonal light scattering it was shown that lymphocytes from a patient with T gamma lymphocytosis were abnormal. Orthogonal light scattering also showed an increase in cytotoxic lymphocytes in a patient with mononucleosis infectiosa and in a splenectomized patient. Orthogonal light scattering of lymphocyte subpopulations showed that the leu8+ population of a patient with mononucleosis infectiosa was bidisperse. For elderly donors the occurrence of CD3+, CD4+, CD8+, and HNK-1+ lymphocytes with a large orthogonal light scattering varied considerably. The CD8+ lymphocytes of these donors consisted mainly of cytotoxic lymphocytes. These results show that determination of light-scattering properties of lymphocytes may yield important diagnostic information and can indicate when further investigation of the lymphocytes by means of immunofluorescence is necessary.     

The requirement for cell surface galactosyl residues during oxidative activation of normal and chronic lymphocytic leukemia lymphocytes.

Galactose oxidase stimulated normal and leukemic lymphocytes to undergo DNA synthesis and cell division. Although the response of normal lymphocytes to galactose oxidase was enhanced with neuraminidase pretreatment, substantial activation of leukemic lymphocytes required pretreatment with neuraminidase. Leukemic lymphocytes exhibited maximal response to neuraminidase-galactose oxidase later than that observed in normal lymphocytes. Treatment of lymphocytes with trypsin diminished their response to galactose oxidase. When lymphocytes were pretreated with β-galactosidase to specifically remove cell surface galactosyl residues, the response to galactose oxidase was prevented. The response of normal and leukemic lymphocytes to sodium periodate was also reduced after treatment with galactose oxidase. These data support the concept that oxidation of cell surface galactosyl residues is critical during lymphocyte activation.     

Distinctive functional properties of human blood L lymphocytes: a comparison with T lymphocytes, B lymphocytes, and monocytes.

Human blood lymphocytes with high affinity Fc receptors have been operationally named L lymphocytes because of membrane-labile IgG markers. L lymphocytes lack membrane-incorporated immunoglobulin and do not form rosettes with sheep red blood cells coated with IgM antibody and mouse complement. These lymphocytes are capable of binding IgG in normal human serum at 4 degrees C and will form rosettes with human lymphocytes coated with Ripley IgG. In this study, functional in vitro properties of isolated L lymphocytes were compared with T lymphocytes, B lymphocytes, and monocytes. To obtain these mononuclear populations, first, plastic adherent monocytes were harvested. T lymphocytes were then isolated by centrifugation of E rosette-forming cells, and other rosetting techniques were employed to isolate L and B lymphocytes by negative selection. The functional properties of L lumphocytes were completely unlike those of T cells, B cells, or monocytes. L lymphocytes did not proliferate in response to mitogens, soluble antigens, or cell surface antigens. Moreover, this population could not replace monocytes in helping T lymphocytes respond to concanavalin A and pokeweed mitogen. Once T cells were supplemented with monocytes, however, the addition of L lymphocytes to the culture greatly enhanced the T lymphocytes proliferative response to phytohemagglutinin, concanavalinA, purified protein derivative (PPD), and streptokinase/streptodornase. L lymphocytes were not a subset of B cells. They did not spontaneously develop surface Ig in culture, and pokeweek mitogen could not induce them to transform and generate cytoplasmic Ig detectable by immunofluorescence. Mixtures of B cells and T cells responded to pokeweed mitogen better than do T cells alone. In contrast, enhanced reactivity with L and T cell combinations was not observed. Another sharp difference between these two populations was the stimulator capacity of each in mixed lymphocyte culture. When B and L lymphocytes were carefully monocyte-depleted, only B cells were effective stimulators of autologous and allogeneic lymphocytes. In comparison with T cells, B cells, and monocytes, L lymphocytes were the only effective killers of human blood lymphocytes sensitized with IgG. L lymphocytes, then, have cytotoxic potential, but cannot proliferate in response to various stimulants or become antibody-producing cells. These findings suggest that L lymphocytes comprise a third lymphocyte population.     

Quantitative parameters of the IgG binding by Fc receptors and the functional activity of blood lymphocytes in healthy cattle and cattle with chronic lympholeukemia

The binding of IgG to lymphocyte Fc receptors in the blood of healthy cattle and of cattle with chronic lympholeukemia has been studied by fluorometric techniques before and after the incubation of lymphocytes in a serum-free medium at 37 degrees C. The study has shown that changes in the intensity of binding of IgG to Fc receptors of normal and leukemic lymphocytes correlate with changes in the cell-mediated cytotoxic activity of the corresponding lymphocytes. Lower cell-mediated cytotoxic activity of leukemic lymphocytes in comparison with that of normal lymphocytes was parallelled by a lower association constant of IgG with leukemic lymphocytes Fc receptors.     

Interactions between human lymphocytes and paramyxovirus-infected cells: adsorption and cytotoxicity.

The capacity of human lymphocytes to adhere to paramyxovirus-infected monolayers and their capacity to kill paramyxovirus-infected cells was investigated. A large fraction of human lymphocytes was found to adhere firmly to the paramyxovirus-infected monolayers. Predsorption of lymphocytes on mumps virus-infected cells impaired their adsorption to a second cell monolayer of the same type. The cytotoxic activity of lymphocytes against mumps virus-infected cells was also reduced after predsorption on mumps virus- or Newcastle disease virus-infected (NDV) cell monolayers. Exposure of lymphocytes to trypsin did not significantly decrease either adsorption or cytotoxicity. Pretreatment of lymphocytes with neuraminidase (NANase) partly inhibited adsorption whereas cytotoxicity was not decreased. Cell fractionation experiments after rosetting of the lymphocytes with sheep erythrocytes (E) indicated that T cells were equally or better adsorbed than "non-T" cells. Taken together with previous experiments which showed that the majority of T lymphocytes are not cytotoxic against mumps virus-infected cells these results suggest that adherence of lymphocytes to infected cells and cytotoxicity may be unrelated phenomena.     

Human lymphocyte-mouse myeloma somatic cell hybrids: selective hybrid formation.

Fusion of unfractionated human lymphocytes with mouse myeloma cells resulted in proliferating hybrid colonies, almost all producting human Ig. We examined whether this high frequency of Ig production was the result of selective formation of human B lymphocyte-mouse myeloma hybrids, rather than induction of Ig genes in T lymphocytes. Unfractionated peripheral lymphocytes and B lymphocytes from patients with the common variable form of agammaglobulinemia formed proliferating somatic cell hybrid colonies. In contrast, peripheral lymphocytes from a patient with agammaglobulinema who lacked B lymphocytes, as well as albumin gradient fractions of peripheral blood which do not contain B lymphocytes, failed to produce somatic cell hybrids with three different myeloma parent cell lines. B, T, and precursor lymphocytes all had Sendai virus receptors, as witnessed by viral agglutination. We conclude that fusion of human lymphocytes with mouse myeloma cells results in selective hybrid formation, rather than activation of Ig genes in disparate cell types. Only B lymphocyte-mouse myeloma heterokaryons form hybrid cells.     

Quantitative study of deoxycytidine incorporation in large and small lymphocytes of the mouse

Using radioautographic smear preparations of thymocytes and mesenteric lymph node (MLN) cells labelled with three different tritiated pyrimidine deoxyribonucleosides, the incorporation of DNA precursors was studied separately on large lymphocytes and small lymphocytes. Radioautographic reaction due to generally tritiated deoxycytidine ( [G-3H]CdR) labelling in vivo in large lymphocytes was more intense than that in small lymphocytes. When mice were sacrificed 6 hr after the administration of tritiated thymidine ( [3H]TdR), small lymphocytes were labelled more heavily than large lymphocytes. However, labelling intensity with [3H]TdR in large lymphocytes was greatly enhanced by the administration of 5-fluoro-deoxyuridine, whereas in small lymphocytes labelling intensity was only fairly enhanced by the same treatment. When cells were incubated in vitro with 5-tritium labelled deoxycytidine [( 5-3H]CdR) for 10 min, there was no significant difference in labelling intensities between large and small lymphocytes. In the case of [G-3H]CdR incorporation, the labelling intensity in large lymphocytes was found to be significantly stronger than that in small lymphocytes. Large as well as small lymphocytes incorporated [3H]TdR very well in vitro. However, addition of 5 X 0 X 10(-5) M of non-radioactive CdR to the medium greatly decreased the incorporation of [3H]TdR by large lymphocytes, whereas the effect of non-radioactive CdR in small lymphocytes was not so marked as that in large lymphocytes. Furthermore, the [3H]TdR-labelling percentages were decreased at the same rate by the addition of non-radioactive CdR in both large and small lymphocytes. These results indicate that large lymphocytes and a proportion of small lymphocytes have a strong tendency to convert CdR to thymidine mono-phosphate, which is utilized for DNA synthesis, whereas this ability is relatively weak in the rest of small lymphocytes. Thus, it is probably that this metabolic ability changes during the transition of the large lymphocyte to the small lymphocyte.     

The chronic lymphatic leukemia lymphocyte: Correlation of functional,metabolic, and surface immunoglobulin characteristics

This study determined the correlation between the functional capacity of chronic lymphatic leukemia lymphocytes as determined by their response to nonspecific mitogens with their glucose metabolism and surface immunoglobulin characteristics. A majority of patients (12) were found to have lymphocytes with impaired transformation to both PHA and pokeweed mitogens. These cells also had impaired glucose metabolism in unstimulated cultures and failed to have the striking increase in glucose metabolism in response to mitogens which is characteristic of normal lymphocytes. Most of these lymphocytes had IgM surface immunoglobulins. However, we were not able to demonstrate surface immunoglobulins on the lymphocytes of one patient in this group. Two patients were found to have lymphocytes with normal lymphoblastic transformation to PHA and impaired transformation to pokeweed suggesting cells of T origin. The glucose metabolism of these lymphocytes were less impaired in unstimulated cultures than those of the other patients and had a striking increment in glucose metabolism in response to PHA similar to normal lymphocytes. Unexpectedly, these lymphocytes were found to have IgG on their surface suggesting cells of B origin. These results indicate that there may be two groups of CLL patients with clinically similar disease in whom the functional and metabolic characteristics of the lymphocytes are distinct and that the surface immunoglobulin characteristic of lymphocytes may not always predict their functional characteristic.     

Preferential Binding of Mouse Mammary Tumor Virus to B Lymphocytes

Mouse mammary tumor virus (MMTV) has been shown to preferentially infect B lymphocytes in vivo. We have used recombinant envelope-coated fluospheres and highly purified MMTV particles to study the distribution of the viral receptors on fresh mouse lymphocytes. A preferential dose-dependent binding to B lymphocytes was observed which could be competed with neutralizing antibodies. In contrast, T-lymphocyte binding remained at background levels. These results strongly suggest a higher density of viral receptor molecules on B lymphocytes than on T lymphocytes and correlate with the preferential initial infection of B lymphocytes observed in vivo.     

In vitro induction of suppressor cells by plasma of rats with Trypanosoma brucei rhodesiense infection

Mitogenic responses of B and T lymphocytes from spleens of rats infected with Trypanosoma brucei rhodesiense were suppressed. Plasma from infected rats suppressed the mitogenic responses of B and T lymphocytes from spleens of normal uninfected rats. Removal of immune complexes from plasma of infected rats significantly reduced the suppressive effect of the plasma on splenic lymphocytes of normal uninfected rats. Normal thymus cells treated with plasma from infected rats and added to cultures of normal spleen lymphocytes inhibited the mitogenic responses of B and T lymphocytes. We suggest that the interaction of immune complexes and Fc or C3b receptors of T lymphocytes resulted in the in vitro induction or activation of T suppressor lymphocytes.     

A possible association between CFA (I) fimbriae and K99 fimbriae on Escherichia coli and bacterial adherence to human lymphocytes

We have explored a possible association between Escherichia coli binding to human lymphocytes and plasmid coded fimbriae on the bacterial surface. E. coli with or without the plasmid coded membrane CFA(I), K99 and K88 were mixed with freshly-drawn human peripheral blood lymphocytes. When the lymphocytes were mixed with E. coli possessing the CFA(I) fimbriae, 59% of the lymphocytes bound bacteria onto the surface, whereas only 22% of the lymphocytes bound the CFA(I)- derivative. The lymphocytes bound 53% and 56% of two K9+ strains, whereas 22% and 8% of the lymphocytes adhered the same strains without the K99 fimbriae. Twelve per cent and 7% of lymphocytes bound bacteria when the strain was K88+ or K88-, respectively. Likewise a low (8%) adherence to lymphocytes was found when the E. coli did not possess fimbriae or flagella.     

Macrophage-lymphocyte clusters in the immune response to soluble protein antigen in vitro. IV. Synergy of lymphocytes in the formation of clusters.

T-lymphocyte-enriched lymph node lymphocytes from guinea pigs immunized with Mycobacterium tuberculosis produce clusters with macrophages when cultivated on monolayers of syngeneic purified protein derivative of tuberculin (PPD)-pulsed peritoneal macrophages. The clusters consist of a macrophage with a central lymphocyte attached to it, and several peripheral lymphocytes attached to the central one. By mechanical manipulation immune lymphocytes incubated on monolayers of PPD-pulsed macrophages were separated into those which adhered firmly to the macrophages after 4 hr of culture and those which did not adhere. While neither of the two populations was able to produce significant numbers of clusters alone, they did so in combination. The number of macrophage-lymphocyte clusters which are produced in a culture depends not only on the absolute number of immune lymphocytes in the culture, but also on the concentration of lymphocytes per area of the macrophage monolayer, with high concentrations resulting in high numbers of clusters. Autoradiographic studies showed that the DNA-synthesizing lymphocytes physically associated with macrophages were located mainly inside the clusters in cultures with high concentrations of lymphocytes but mainly outside the clusters in cultures with low concentrations of lymphocytes.     

The entry of T and B lymphocytes into rat popliteal lymph nodes undergoing a graft-versus-host reaction

The entry of radiolabeled blood-borne T and B lymphocytes into resting popliteal lymph nodes and popliteal lymph nodes stimulated with semiallogeneic lymphocytes was investigated in rats. Thoracic duct lymphocytes separated into T- and B-lymphocyte populations on nylon-wool columns were radiolabeled with 51chromium and equal numbers of T or B lymphocytes were injected intravenously. While the ratio of T and B lymphocytes in the blood is approximately 3:1 it was found that the ratio of T to B lymphocytes migrating into lymph nodes was approximately 9 T to 1 B lymphocyte in both resting and antigenically stimulated lymph nodes. Since the ratio of T to B lymphocytes in thoracic duct lymph is similar to that of blood, there is a disparity between the number of T cells entering and leaving lymph nodes. These results suggest that some T lymphocytes may return to the blood directly and/or there is increased T lymphocyte death in lymph nodes.     

Autologous mixes lymphocyte reaction in human cord blood lymphocytes: decreased generation of helper and cytotoxic T-cell functions and increased proliferative response and induction of suppressor T cells

T lymphocytes from neonates proliferated significantly more than peripheral blood T lymphocytes from adults in autologous mixed lymphocyte reactions (AMLR). AMLR-activated cord, as compared to adult T lymphocytes, exerted significantly less nonspecific cytotoxic activity on PHA-stimulated adult mononuclear cells and Epstein-Barr virus-transformed target cells. The impaired generation of cytotoxicity of cord T cells was not corrected by Interleukin-2. Blood T lymphocytes from adults activated in AMLR synthesized a helper factor that supported PWM-induced proliferation and immunoglobulin production in both adult and cord B lymphocytes. In contrast, cord blood T lymphocytes failed to produce the helper factor for B lymphocytes. T cells from AMLR cultures established with neonatal lymphocytes showed suppressor activity, as assessed in PWM-stimulated immunoglobulin synthesis of adult peripheral-blood mononuclear cells, significantly higher than that exhibited by T cells from AMLR cultures performed with lymphocytes from adults. Finally, neonatal B lymphocytes could be activated to the production of IgM but not IgG by either adult AMLR-derived helper factor plus PWM or by Epstein-Barr virus, whereas adult B cells secreted both IgM and IgG under the same type of stimulation.     

Morphological Differences between Thymus- and Bone Marrow-Derived Lymphocytes

Simple morphological criteria are described, enabling one to distinguish two types of lymphocytes in normal unstimulated mice: 'Th', predominant in the thymus (90-92%) and 'Bm', predominant in the bone-marrow (85-93%). Their distribution in the peripheral lymphoid organs agreed significantly with the observed average distribution of T and B lymphocytes. Thy-mectomized and irradiated bone-marrow-restored CBA mice, as well as nude mice, showed a striking lack of Th cells. In vitro depletion of θ-bearing or Ig-bearing lymphocytes lead to a correlated depletion of Th or Bm lymphocytes, respectively: blood lymphocytes treated with anti-θ serum + complement showed a decrease in Th lymphocytes corresponding to the percentage of cells killed by the anti-θ serum. The selective retention of Ig-bearing spleen cells on Wigzell's anti-lg columns caused a depletion of Bm lymphocytes. It is concluded that the Th type is the morphological expression of the T lymphocytes, and the Bm type that of the B lymphocytes. The implications of this way of distinguishing T and B lymphocytes by simple morphology, and of the actual relationships between T and B cell differentiation, are discussed.     

Decidual lymphocytes of human spontaneous abortions induce apoptosis but not necrosis in JEG-3 extravillous trophoblast cells

The human decidua contains an unusually high proportion of lymphocytes, mainly NK and T cells, which are potentially cytotoxic to the trophoblast when they are stimulated with certain cytokines. Given the high incidence of spontaneous abortion in humans and other species, our working hypothesis is that decidual lymphocytes are involved in immunological mechanisms that attack the trophoblast and induce abortion when any gestational problem arises. To test this hypothesis, flow cytometry was used to compare decidual lymphocyte populations in first-trimester spontaneous abortions and elective terminations of first-trimester pregnancy. We found significantly higher proportions of decidual lymphocytes that expressed activation markers, and of T cells (mainly T helper cells) in spontaneous abortions than in elective terminations of pregnancy. Decidual lymphocytes from spontaneous abortion, like decidual lymphocytes from elective termination of pregnancy and peripheral blood lymphocytes, were however, unable to lyse the JEG-3 extravillous cytotrophoblast cell line in a (51)Cr-release assay. Nevertheless, decidual lymphocytes from spontaneous abortion, unlike decidual lymphocytes from elective termination of pregnancy and peripheral blood lymphocytes, induced apoptosis in JEG-3 cells as determined by DNA fragment-release assay. Hematoxylin and eosin staining showed a significantly higher proportion of apoptotic JEG-3 cells when these cells were treated with decidual lymphocytes from spontaneous abortion than when JEG-3 cells were cultured with decidual lymphocytes from elective termination of pregnancy. The ultrastructural signs of apoptosis were confirmed by electron microscopy. These data support the hypothesis that activated decidual lymphocytes participate in human spontaneous abortion by inducing apoptosis but not necrosis of the trophoblast.