Changes in plasma membrane phospholipids inhibit antibody-mediated lysis

A variety of mechanisms have been proposed to explain how tumors evade immune destruction. This work has identified one such mechanism that determines susceptibility to immune lysis; membrane phospholipid composition altered susceptibility to antibody plus complement (Ab+C)-mediated lysis. Effects on antibody plus complement-mediated lysis were correlated with levels of major histocompatibility complex (MHC) molecules but not inherent resistance to complement damage. This cellular mechanism could be a means by which tumor cells escape immune detection and destruction.     

Interactions between immature porcine Leydig and Sertoli cells in vitro

Summary Interactions between Leydig and Sertoli cells, as well as a stimulatory effect of FSH on Leydig cell activity, have been reported in many studies. In order to investigate these interactions, the ultrastructure of immature pig Leydig cells under different culture conditions has been studied. When cultured alone in a chemically defined medium, there is a marked regression of the Leydig cell smooth endoplasmic reticulum and a swelling of the mitochondria. Addition of FSH or hCG does not prevent these phenomena. Co-culturing of Leydig cells with Sertoli cells from the same animal maintains the smooth endoplasmic reticulum at the level seen in vivo and in freshly isolated Leydig cells. The addition of FSH to the co-culture stimulates its development and increases Leydig cell activity, as assessed by an increase in hCG binding sites and an increased steroidogenic response to hCG. These results suggest that Sertoli cells exert a trophic effect on Leydig cells, and that the stimulatory effect of FSH on Leydig cell function is mediated via the Sertoli cells. These results reinforce the concept of a local regulatory control of Leydig cell steroidogenesis.Post-Doctoral fellow supported by CIRIT, Generalitat de Catalunya, Spain     

Oncogenicity of human papillomavirus- or adenovirus-transformed cells correlates with resistance to lysis by natural killer cells.

The reasons for the dissimilar oncogenicities of human adenoviruses and human papillomaviruses (HPV) in humans are unknown but may relate to differences in the capacities of the E1A and E7 proteins to target cells for rejection by the host natural killer (NK) cell response. As one test of this hypothesis, we compared the abilities of E1A- and E7-expressing human fibroblastic or keratinocyte-derived human cells to be selectively killed by either unstimulated or interferon (IFN)-activated NK cells. Cells expressing the E1A oncoprotein were selectively killed by unstimulated NK cells, while the same parental cells but expressing the HPV type 16 (HPV-16) or HPV-18 E7 oncoprotein were resistant to NK cell lysis. The ability of IFN-activated NK cells to selectively kill virally transformed cells depends on IFN's ability to induce resistance to NK cell lysis in normal (i.e., non-viral oncogene-expressing) but not virally transformed cells. E1A blocked IFN's induction of cytolytic resistance, resulting in the selective lysis of adenovirus-transformed cells by IFN-activated NK cells. The extent of IFN-induced NK cell killing of E1A-expressing cells was proportional to the level of E1A expression and correlated with the ability of E1A to block IFN-stimulated gene expression in target cells. In contrast, E7 blocked neither IFN-stimulated gene expression nor IFN's induction of cytolytic resistance, thereby precluding the selective lysis of HPV-transformed cells by IFN-activated NK cells. In conclusion, E1A expression marks cells for destruction by the host NK cell response, whereas the E7 oncoprotein lacks this activity.     

Modulation of natural killer-mediated lysis by red blood cells

Natural killer (NK)-mediated cytotoxicity is significantly enhanced in the presence of red blood cells (RBC). The enhancement is dose dependent on the number of RBC present and can be induced by autochthonous, allogeneic, or xenogeneic RBC. The enhancement was demonstrated in cytotoxicity against two different NK-sensitive tumor target cell lines, K562 and U937, and by three different assay systems, chromium release, lactate dehydrogenase release, and inhibition of thymidine incorporation. RBC directly enhance the cytotoxic activity of NKH-1/Leu19+ large granular lymphocyte NK cells. Intact RBC have to be present during the cytotoxicity assay to induce the enhancement, which probably occurs at a postbinding, preprogramming phase. The anti-CD2 antibody Leu5 cannot block the enhancement at a concentration inhibitory to lymphocyte rosetting with sheep RBC, suggesting that the enhancement is not induced by interaction through the CD2 antigen. These results indicate that RBC are a potent modulator of NK cytotoxicity and suggest that in vitro NK studies using purified lymphocytes may not always truly reflect NK activity in the blood stream.     

Multinucleate Sertoli cells in aged human testis

Summary The present investigation documents morphological characteristics of human Sertoli cells of aged males. Testicular material was obtained from 35 patients (age 62–84 years) with carcinoma of the prostate who had received no previous anticancer therapy. As revealed by light and electron microscopy the appearance of the germinal epithelium showed great individual variations. In all cases examined, however, the occurrence of multinucleate Sertoli cells was a common finding. In seminiferous tubules with intact spermatogenesis these cells closely resembled the normally occurring variants, whereas they displayed features reminiscent of immaturity in the absence of germ cells. It is hypothesized that the nuclei of Sertoli cells in the special situation of aging may resume the capacity to divide, an ability normally restricted to immature cells. Thus, mitosis without subsequent cytokinesis might be an explanation for the formation of multinucleate Sertoli cells.The authors wish to thank Dr. R. Hubmann, Hamburg, for kindly supplying the testicular material. The excellent technical assistance of Mrs. E. Roosen-Runge, Mrs. E. Schäfer, and Mrs. A. Stromeyer is gratefully acknowledgedSupported by grants from the Deutsche ForschungsgemeinschaftPresented in part at the Kleinkonferenz der Deutschen Forschungsgemeinschaft, Schloß Auel, October 3–5, 1980     

Free fatty acids inhibit cytotoxic T lymphocyte-mediated lysis of allogeneic target cells

Short term exposure of murine CTL clones to long chain cis unsaturated free fatty acid (FFA) inhibits alloantigen specific lysis of cognate target cells, whereas long-chain saturated FFA have no effect. Inhibition of lysis occurs when cis FFA is added before or within 10 min after CTL-target cell conjugate formation and thus appears to interfere with lethal hit delivery. Our previous studies have shown that similar treatment with cis FFA inhibits, in CTL, the Ag stimulated increase in intracellular calcium and degranulation, suggesting that inhibition of lysis probably results from perturbation of the CTL signaling pathway. However, inhibition of lysis is probably not due to the inhibition of the rise in intracellular calcium or degranulation, because lysis can occur under conditions in which FFA inhibit degranulation and because cis FFA inhibit calcium-independent killing. Inhibition of lysis is detectable at unbound FFA concentrations less than 1 microM and is generally complete at concentrations less than 5 microM. Although these levels of FFA are somewhat higher than reported for normal physiologic conditions, plasma FFA levels can be elevated into this range in states of stress and disease, suggesting that FFA modulation of the immune response has important physiologic consequences.     

Systematic comparison of antibody-mediated mechanisms of keratinocyte lysis in vitro

We have conducted a systematic comparison of lysis of TNP-coated keratinocyte targets by TNP-specific antibody, by antibody plus complement, by antibody-dependent cellular cytotoxicity (ADCC), and by natural killing with the use of monocyte, lymphocyte, and neutrophil effectors. With chromium-release assays, human keratinocytes, HEp-2 cells (transformed human keratinocytes), PAM 212 cells (transformed mouse keratinocytes), and RSC (transformed rabbit keratinocytes) were all susceptible to monocyte- and lymphocyte-mediated ADCC (p less than 0.01 to p less than 0.02). All trypsinized keratinocyte targets were also susceptible to natural killing by monocyte or lymphocyte effectors (p = 0.05 to p less than 0.001). Antibody and antibody plus complement were poor mediators of keratinocyte lysis. If protein and complex lipid synthesis of keratinocytes were inhibited by 16-hr cycloheximide preincubation, then keratinocytes were susceptible to complement-mediated lysis, implying that the resistance of these cells to complement may be due to repair of transmembrane pores. Comparison of chromium-release assays with fluorescein diacetate dye uptake viability assays showed that human keratinocytes were still susceptible to monocyte and lymphocyte ADCC but not to antibody, antibody plus complement, or natural killing. The reproducible and uniform susceptibility of normal and transformed keratinocyte targets from three different species to monocyte and lymphocyte ADCC supports the hypothesis that this mechanism of cellular lysis may be important in antibody-associated diseases of epidermal cytotoxicity.     

Effect of catecholamines on porcine Sertoli and Leydig cells in primary culture

The accumulation by purified immature porcine Leydig and Sertoli cells of cyclic adenosine 3',5'-monophosphate in the presence of 1-methyl-3-isobuthylxathine was studied and their respective testosterone and 17 beta-estradiol production in response to catecholamines was assessed in vitro. These substances increased both basal and FSH-stimulated cyclic adenosine 3',5'-monophosphate accumulation in Sertoli cells. In contrast, catecholamines slightly enhanced basal cyclic adenosine 3',5'-monophosphate production but inhibited its human chorionic gonadotropin-stimulated accumulation by Leydig cells. Catecholamines had no effect on basal and stimulated testosterone release by these cells, while dopamine inhibited 17 beta-estradiol synthesis by Sertoli cells. Using various alpha- and beta-adrenergic agonists and antagonists, beta-receptors, likely of the beta 1-subtype, were shown to be present in both cell lines. Taken together these data suggest the presence of a cyclic adenosine 3',5'-monophosphate-linked adrenergic receptor in porcine Leydig and Sertoli cells, the role of which remains to be determined.     

Effect of hyaluronan to inhibit caspase activation in porcine granulosa cells

We studied the ability of hyaluronan (HA) to inhibit apoptosis in porcine granulosa cells. The granulosa layer with cumulus-oocyte complex is cultured in media supplemented with follicle stimulating hormone (FSH) and 4-MU an inhibitor of hyaluronan synthases. The concentration of HA significantly increased after supplemented with FSH, but significantly decreased with 4-MU. CD44, receptor of HA, expressed after cultured with FSH, decreased in addition low concentration of 4-MU, whereas not detected in high concentration of 4-MU, indicating parallel relation between the amount of HA and CD44 expression. The 4-MU treatment also decreased the expression of procaspase-3, -8, -9 suggesting that inhibition of HA synthesis leads to activation of these caspases. Moreover, addition of anti-CD44 antibody decreased the expression of procaspases suggesting that perturbation of HA-CD44 binding leads activation of caspases. Hence, HA has ability to inhibit apoptosis and HA-CD44 binding is important on apoptosis inhibitory mechanism in porcine granulosa cells.     

Fractalkine expression induces endothelial progenitor cell lysis by natural killer cells

Background

Circulating CD34⁺ cells, a population that includes endothelial progenitors, participate in the maintenance of endothelial integrity. Better understanding of the mechanisms that regulate their survival is crucial to improve their regenerative activity in cardiovascular and renal diseases. Chemokine-receptor cross talk is critical in regulating cell homeostasis. We hypothesized that cell surface expression of the chemokine fractalkine (FKN) could target progenitor cell injury by Natural Killer (NK) cells, thereby limiting their availability for vascular repair.

Methodology/Principal Findings

We show that CD34⁺-derived Endothelial Colony Forming Cells (ECFC) can express FKN in response to TNF-α and IFN-γ inflammatory cytokines and that FKN expression by ECFC stimulates NK cell adhesion, NK cell-mediated ECFC lysis and microparticles release in vitro. The specific involvement of membrane FKN in these processes was demonstrated using FKN-transfected ECFC and anti-FKN blocking antibody. FKN expression was also evidenced on circulating CD34⁺ progenitor cells and was detected at higher frequency in kidney transplant recipients, when compared to healthy controls. The proportion of CD34⁺ cells expressing FKN was identified as an independent variable inversely correlated to CD34⁺ progenitor cell count. We further showed that treatment of CD34⁺ circulating cells isolated from adult blood donors with transplant serum or TNF-α/IFN-γ can induce FKN expression.

Conclusions

Our data highlights a novel mechanism by which FKN expression on CD34⁺ progenitor cells may target their NK cell mediated killing and participate to their immune depletion in transplant recipients. Considering the numerous diseased contexts shown to promote FKN expression, our data identify FKN as a hallmark of altered progenitor cell homeostasis with potential implications in better evaluation of vascular repair in patients.     

Diverse multidrug-resistance-modification agents inhibit cytolytic activity of natural killer cells

Multidrug resistance (MDR) is the phenomenon in which cultured tumor cells selected for resistance to one chemotherapeutic agent simultaneously acquire resistance to several apparently unrelated drugs. MDR in tumor cells is associated with the over-expression of P-glycoprotein, an ATP-dependent cell-membrane transport molecule. P-glycoprotein is also expressed in several normal tissues but its physiological role(s) is unknown. We recently observed that a hierarchy of MDR-like activity exists among human peripheral blood lymphocytes in the order CD8>CD4>CD20 (cytoxic/suppressor T cells, helper T cells and B cells respectively). In this study, we report that natural killer (NK) cells also express MDR-like activity. This activity could be inhibited with verapamil or solutol HS-15, two agents that reverse MDR in tumor cells. These, and four additional reversing agents, were used to investigate the possible role of P-glycoprotein in NK cells. We observed that at 10% of their IC₅₀, five of six reversing agents inhibited NK-cell-mediated cytotoxicity; at higher (but non-toxic) doses, all six agents were inhibitory. These data suggest that NK-cell-mediated cytotoxicity may require the functional expression of an efflux molecule similar or identical to P-glycoprotein.This work was supported in part by grants from the National Cancer Institute (CA 57 470), the American Cancer Association — Illinois Division (no. 91-48), The Arthur Andersen Foundation and the Rayman, Freidman, Ditore and Rosenmutter Funds.     

Human choriocarcinoma cell resistance to natural killer lysis due to defective triggering of natural killer cells

The trophoblast, the outermost layer of the human placenta, lacks expression of the classical human leukocyte antigen (HLA) class I molecules. This prevents allorecognition by T cells but raises the question of what protects the trophoblast from natural killer (NK) cells. In a previous study, we have shown that choriocarcinoma cell (CC) resistance to NK lysis was mainly independent of HLA class I molecules. In the present study, we postulated that CC may prevent activation of NK cells by failing to stimulate their triggering receptors (TR). To test this hypothesis, we evaluated the lysis of JAR and JEG-3 CC after effective cross-linking and activation of NK cells by means of lectins or antibodies. Our results show that NK-resistant CC were sensitive to lysis by unstimulated peripheral blood lymphocytes in the presence of phytohemagglutin (PHA), to antibody-dependent cell cytotoxicity in presence of anti-Tja antibodies, and to monoclonal antibody redirected killing using anti-TR antibodies anti-CD16 and anti-CD244/2B4. Finally, CC fail to express CD48, the ligand for CD244/2B4. These results indicate that the resistance of CC to lysis results primarily from defective NK cell activation, at least partially due to the lack of expression of ligands, such as CD48, involved in the triggering of NK cells.     

Catecholestrogens inhibit basal and luteinizing hormone-stimulated androgen production by porcine thecal cells

It has been shown recently that catecholestrogens are produced by cultured porcine granulosa and thecal cells, and that they influence porcine granulosa cell steroidogenesis in a similar manner to estradiol-17 beta (E2). The present studies were performed to determine if catecholestrogens also play a role in the regulation of porcine thecal cell steroidogenesis and to compare their actions to those of E2. Thecal cells were obtained from prepubertal gilts and cultured in a serum-free medium for 48 h. Thecal cell androstenedione production under basal and luteinizing hormone (LH)-stimulated conditions was significantly inhibited by adding E2 or catecholestrogens to the culture medium. Treatment of basal and LH-stimulated cultures with increasing concentrations of E2 or catecholestrogens (0.1-10 micrograms/ml) caused a dose-and time-dependent inhibition of androstenedione production. The inhibitory effect of the catecholestrogens, but not of E2, was enhanced when the cultures contained the catechol-O-methyl transferase inhibitor, U-0521. Studies to determine the mechanism(s) of action of the catecholestrogens showed that E2 and catecholestrogen actions are exerted at a site(s) distal to cyclic adenosine 3'5' monophosphate (cyclic AMP) generation, because neither agent affected the basal or LH-stimulated accumulation of extracellular cyclic AMP, while causing a significant inhibition of androstenedione production. E2 or catecholestrogen treatment also inhibited androstenedione production stimulated by prostaglandin E2 and dibutyryl cyclic AMP. In addition, both E2 and catecholestrogen treatment significantly decreased basal and LH-stimulated 17 alpha-hydroxyprogesterone production, while significantly increasing pregnenolone production. Progesterone production in the presence of E2 or catecholestrogens showed small but statistically insignificant increases.(ABSTRACT TRUNCATED AT 250 WORDS)     

Rapid kinetics of lysis in human natural cell-mediated cytotoxicity: Some implications

The entire lytic process of natural cell-mediated cytotoxicity against sensitive target cells can occur rapidly, within minutes. This was demonstrated by ⁵¹chromium release and in single-cell assays. At the cellular level, most of the target cell lysis occurred within 15–30 min after binding to effector cells. The enriched natural killer cell subpopulation of lymphocytes obtained by Percoll density gradient centrifugation (containing >70% large granular lymphocytes (LGL)) was the most rapidly lytic population by ⁵¹chromium release. However, in the single-cell assay, the rate of lysis of bound target cells was quite similar for the LGL-enriched effector subpopulation and the higher density subpopulation of effector cells recognized previously. Both the light and dense effector cells contained similar numbers of target binding cells. Therefore, that the light subpopulation effected lysis more rapidly and to a greater extent than the dense subpopulation suggested that the low-density effector cells probably recycled more rapidly than those of higher density. This was corroborated by the finding that when conjugates were formed at 29 °C for the single-cell assay, a significant number of dead unconjugated targets could be observed only on the slides made with the LGL-enriched effector cells but not on those made with dense effector cell. Lysis continued to increase in the chromium-release assay probably because of recycling, recruitment, and/or heterogeneity of the effector cells, and/or because of heterogeneity or delayed death of the target cells.     

GD3-reactive antibodies can inhibit the lysis of autologous tumor cells by tumor-infiltrating lymphocytes

Summary GD3 is expressed in high concentrations on melanoma cells and may serve as a useful target antigen for mAb-mediated immunotherapy. Monoclonal antibodies (mAbs) against GD3 stimulate cell-mediated immune responses against tumor cells in vitro and this activity may contribute to antitumor effects in patients with melanoma treated with GD3-reactive mAbs. In the present study the effects of GD3-reactive mAbs on autologous tumor cell lysis by a human melanoma-derived tumor-infiltrating lymphocyte (TIL) population were examined. Unlike results reported for other GD3⁺ T cells isolated from melanoma patients, the tumor-specific lytic activity of the TIL line was inhibited by incubation with mAbs against GD3. Other melanoma-reactive mAbs, including those against GD2 and the high-molecular-weight melanoma-associated Ag, had no effect on the TIL lytic activity. Overall, these results indicate that mAbs against GD3 may have different effects on T cell/tumor cell interactions.     

Spent media from immature seminiferous tubules and Sertoli cells inhibit adult rat Leydig cell aromatase activity

Adult rat Leydig cell aromatase activity is stimulated 2.5 fold by LH or dbcAMP. Spent media prepared from seminiferous tubules or Sertoli cells of immature rats depress both the basal and the LH stimulated estradiol syntheses (25 and 20% decreases, respectively). These inhibitory effects are further enhanced when FSH is added to the culture medium of seminiferous tubules or Sertoli cells. Rat serum as well as culture media from other cell lines are ineffective while seminiferous tubule media from other immature animals (mouse, guinea-pig, calf) inhibit the aromatase activity. This Sertoli cell factor is a heat stable protein (molecular weight greater than 10 kDa), different from the LHRH-like Sertoli cell compound, which acts on the aromatase activity at a step beyond the adenylate cyclase.     

Human alveolar macrophage FcR-mediated cytotoxicity. Heteroantibody- versus conventional antibody-mediated target cell lysis

Human alveolar macrophage have three distinct receptors for IgG: FcRI, FcRII, and FcRIII. In order to compare the ability of these receptors to mediate target cell lysis, three different assay systems were examined. First, we studied lysis of chicken E (CE) opsonized with heteroantibodies, which are synthetic antibodies composed of Fab fragments with anti-FcR activity covalently linked to Fab fragments with anti-CE activity. We found alveolar macrophage readily lysed heteroantibody-opsonized CE via each of the three FcR classes (FcRI, 20 +/- 5%; FcRII, 27 +/- 7%; and FcRIII, 13 +/- 13%, p less than 0.05). Non-FcR-dependent lysis of anti-beta 2-microglobulin x anti-CE heteroantibody-opsonized CE was not detected. Second, lysis of hybridoma cell lines bearing anti-FcR antibodies on their cell surface was examined to assess killing of "tumor-like" target cells. Whereas peripheral blood monocytes and lymphocytes were able to lyse hybridoma cell lines bearing surface anti-FcR mAb, alveolar macrophages were not. Third, activity of alveolar macrophage FcR was examined in a conventional antibody-dependent cellular cytotoxicity assay by using O+ (R1,R2) human RBC opsonized with human anti-D and anti-CD serum as target cells. We found lysis of anti-D and anti-CD opsonized human RBC was mediated exclusively via FcRI. No activity of FcRII or FcRIII was detected in these latter assays even if performed under conditions that impair FcRI activity. Thus, all three FcR present on alveolar macrophage mediate lysis of heteroantibody-opsonized CE; in contrast, with the use of a conventional antibody-dependent cellular cytotoxicity assay, only FcRI activity was detected. We were unable to demonstrate lysis of anti-FcR-bearing hybridoma cell lines by alveolar macrophages.     

Neonatal treatment with naloxone increases the population of Sertoli cells and sperm production in adult rats

Endogenous opioid peptides play an important role in the ontogenesis of the functional and morphological parameters of the seminiferous epithelium. The aim of this study was to evaluate the effects of neonatal manipulations with naloxone, an opioid antagonist, on the population of Sertoli cells and on sperm production in adult rats. Rats were assigned to receive 8 mug per gram of body weight twice a day with interval of 8 h of naloxone and they were compared to a control group receiving saline. Naloxone groups presented the following findings when compared to the control group: increased body weight from the 2nd to the 27th day; a smaller seminiferous epithelium height, smaller seminiferous tubule diameter, increased number of Sertoli cells and daily sperm production per testis, increased daily sperm production per gram per testis and increased total length of the seminiferous tubule of the treated groups. According to our study, the neonatal treatment with naloxone during the critical period of testis development was able to change the proliferative dynamics of Sertoli cells by an intra and/or extra testicular blockage of opioid receptors, confirming the direct relation between the number of Sertoli cells and the number of spermatozoids.     

Recognition and lysis by natural killers of tumor cells with participation of laminin

According to the data obtained in the present work, the receptor complex of mouse natural killers (NK) includes laminin, an antibody that blocks NK activity (NKA) regardless of the presence of a complement. Preincubation of mouse splenocytes with antilaminin serum led to a decrease in their NKA towards tumor cell-stargets (CT) with the NKA activity decreasing by a factor of 2 with respect to cultivated cells of rat hepatoma HTC and by a factor of 10 with respect to cultivated cells of human erythroblastosis K562. Pretreatment of splenocytes with normal nonimmune serum did not lead to a change of NKA. The pattern after tumor-cell preincubation with antilaminin serum was quite different; pretreatment of CT K562 led to a twofold decrease in the sensitivity of these cells to NK lysis, whereas the pretreatment of CT K562, on the contrary, made them twice as sensitive to NK lysis. Electrophoretic separation of proteins of CT plasma membranes with subsequent immunoblotting with antilaminin immune serum revealed the presence of laminin on HTC cell plasma membrane, which was identified as laminin 8/9 by the mass-spectrometry method, while no laminin was detected on K562 cells. Preincubation of splenocytes with laminin did not affect NKA with respect to CT K562 and HTC. Pretreatment of CT K562 and HTC with laminin decreased the NKA to zero. The obtained data allow for the suggestion of the doubtless participation of laminin and its receptors in CT cytolysis by NK.     

Recognition and lysis by natural killers of tumor cells with participation of laminin

According to the data obtained in the present work, the receptor complex of mouse natural killers (NK) includes laminin, antibody to which blocks EK-activity (NKA regardless of the presence of complement. Preincubation of mouse splenocytes with anti-laminin serum led to a decrease in their NKA towards tumor cells-targets (CT), the NKA activity decreasing 2 times with respect to cultivated cells of rat hepatoma HTC, while 10 times - to cultivated cells of human erythroblastosis K562. Pretreatment of aplenocytes with noraml nonimmune serum did not lead to a change of NKA. Quite different was the pattern after the tumor cell preincubation with anti-laminin serum: pretreatment of CT K562 led to a twofold decrease in sensitivity of these cells to NK-lysis, whereas the pretreatment of CT K562, on the contrary, made them twice sensitive to NK-lysis. Electrophoretic separation of protein of CT plasma membranes with subsequent immunoblotting with anti-laminin immune serum revealed the presence oflaminin on HTC cell plasma membrane, which was identified as laminin 8/9 by the mass-spectrometry method, while no laminin was detected on K562 cells. Preincubation of splenocytes with laminin did nor affect NKA with respect to CT K562 and HTC. Pretreatment of CT K562 and HTC with laminin decreased the NKA to zero. The obtained data allow suggesting a doubtless participation of laminin and its receptors in CT cytolysis by NK.