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Decreased natural killer (NK) activity as well as interleukin 2 (IL-2) are risk factors for the progression of cervical carcinoma. NK activity and IL-2 may be thymus controlled. Plasma levels of active thymulin, a zinc-dependent thymic hormone (ZnFTS), are reduced in cancer because of the low peripheral zinc bioavailability. Zinc and thymulin are relevant for normal immune functions. Alpha2-macroglobulin is an inhibitor of matrix metalloproteases (MMPs) against invasive tumour proliferation. Because alpha2-macroglobulin has a binding affinity (Kd) for zinc that is higher than does thymulin, it may play a key role in immune efficiency in cancer. Plasma samples of 22 patients (age range 35-60 years) with locally advanced squamous cervical carcinoma and with FIGO stage Ib2-IIb were examined. They showed reduced active thymulin, decreased NK activity and IL-2 production, increased soluble IL-2 receptor (sIL-2R) and augmented alpha2-macroglobulin in the circulation, whereas plasma zinc levels were within the normal range for age. Significant positive correlations were found between zinc or active thymulin and alpha2-macroglobulin (r = 0.75, P < 0.01, r = 0.78, P < 0.01, respectively) in cancer patients. In vitro zinc increases IL-2 production from peripheral blood mononuclear cells (PBMCs) of cancer patients. These data suggest that an increase in alpha2-macroglobulin, which competes with thymulin for zinc binding, may be involved in causing a thymulin deficit with a consequent decrease of IL-2 and NK cytotoxicity. Thus, physiological zinc treatment in cervical carcinoma maybe restores impaired central and peripheral immune efficiency.

In this study we evaluated the effect of long-term melatonin (MEL) treatment on the cytotoxic activity and number of natural killer (NK) cells and the proliferative response of spleen lymphocytes to phytohemagglutinin (PHA) or interleukin-2 (IL-2) in old mice. Seventeen-eighteen month-old Balb/c mice were supplemented with MEL (40-50 microg/day/mouse) and sacrificed after eight months. The MEL supplementation was unable to recover the low levels of both endogenous and IL-2-induced NK cell activity found in old untreated mice. Also the NK cell number was unaffected by MEL treatment. The spleen lymphocyte proliferative response to both PHA and IL-2 was not different in old MEL-treated compared to old untreated mice. These results indicate that long-term MEL supplementation does not recover the age-related deterioration of NK cell activity and lymphocyte proliferative capacity.

In a previous work we demonstrated that chronic in vivo antalgic therapy of cancer patients with morphine reduced the endogenous cytotoxic activity of natural killer (NK) cells, while increasing the development of lymphokine activated killer (LAK) cell cytotoxicity. In order to investigate the mechanisms by which morphine affects NK and LAK cell function further, we evaluated the modulation exerted by short- or long-term morphine administration on either NK/LAK cell cytotoxicities or plasma levels of prolactin (PRL) and other immunomodulating neurohormones. An intravenous morphine injection (10 mg) significantly increased the plasma levels of PRL, reduced the cytotoxic activity of NK cells, and increased the development of LAK cell activity 30 min after drug injection in neoplastic patients. The administration of bromocriptine before the injection of morphine prevented both PRL augmentation and the increase in LAK cell activation, although it did not prevent the inhibition of NK cytotoxicity. The chronic oral administration of morphine (90 +/- 30 mg/day for 1 month) also resulted in higher PRL levels; the NK and LAK cell activities were, respectively, lower than or higher than those found in neoplastic patients untreated with morphine. The plasma levels of thyrotropin (TSH), adrenocorticotropic hormone (ACTH) and cortisol were not significantly modified in either short- or long-term experiments. The absolute number and the percentages of lymphocyte populations, as well as the percentage of IL-2 receptors, were not modified after short-term morphine administration whereas little changes of T lymphocyte populations and NK cell number were observed after oral treatment with morphine. In vitro morphine did not affect the development of LAK cell activity. In conclusion, our findings indicate that morphine reduces NK cytotoxicity and increases the development of LAK cell cytotoxicity after short- and long-term administration. The effect of morphine on LAK cell activation but not on NK cell reduction is related to the modulation of PRL levels determined by the opioid drug.

Objective: To investigate the effects of pharmacologic suppression of ovarian function on the immune system, with respect to the clinical outcome of endometriosis and the possibility of an immunoendocrine combined treatment., Methods: After informed consent, 25 of 37 patients with revised American Fertility Society stage III and IV endometriosis who underwent postoperative medical treatment were selected and enrolled for this immunoendocrine longitudinal study. Medical treatment consisted of tryptorelinum depot injection, 3.75 mg/month for 24 weeks. Blood samples were collected before the first injection in the early follicular phase, day 2-3 of the cycle, and during medical treatment (every 4 weeks) and follow-up (every 6 months). At the end of the study, we had ten blood samples per patient to evaluate the cytotoxic activity, the number of natural killer cells, and the serum levels of estradiol. Natural killer activity was determined against the K562 cell line by target cell retention of the fluorescent dye carboxyfluorescein diacetate., Results: A positive immunomodulating effect was observed during GnRH agonist administration. In particular, a significant progressive increase in natural killer cell activity was defined within the first 12 weeks of medical treatment; after three injections, we observed the highest values of cytotoxicity, with a median of 7.1 lytic units (range 0.3-14.0; P = .02). Natural cytotoxicity then decreased toward a plateau, which persisted during therapy completation and follow-up, with slight fluctuations. In patients who had recurrence, the values of natural killer cell activity were constantly lower than those in patients with disease-free follow-up, particularly within the first 12 weeks of medical treatment.

Endometriosis is an estrogen-dependent disease affecting women during their reproductive years. An abnormal immune function and, in particular, a decreased natural killer (NK) cell activity have been found in endometriosis, suggesting a role of the immune system in the pathophysiology of the disease. We have recently evidenced a significant inverse relationship between 17-beta-estradiol plasma levels and NK cytotoxicity in endometriosis patients. In this study we have investigated the combined role of 17-beta-estradiol (E2) and prolactin (PRL) in the regulation of NK cell activity during the progression of endometriosis, by evaluating the correlation among E2, PRL, and other immunomodulating neurohormones on both the cytotoxic activity and the number of NK cells in women at different stages of endometriosis. The early stages (I/II) of endometriosis are characterized by increased plasma levels of either E2 or PRL without significant alterations of NK cell activity in comparison with healthy subjects. The progression to advanced stages (III/IV) of the disease is associated with a further increase of E2 levels, a decrease of PRL plasma concentrations (with an increase of E2/PRL ratio), and an impairment of NK cytotoxicity. The plasma levels of both E2 and PRL and the E2/PRL ratio are significantly correlated with the values of NK cytotoxicity in advanced stages of endometriosis. Either the absolute number or the relative percentage of CD16+ or CD56+ peripheral lymphocytes are not significantly different between patients at either stages I/II or III/IV and healthy controls. Plasma levels of progesterone (P) and luteinizing hormone (LH), are not significantly changed in different stages of endometriosis with respect to healthy controls. The significant decrease of follicle-stimulating hormone (FSH) plasma levels found in either stages I/II or III/IV endometriosis patients is not correlated with the NK cell activity. In conclusion, at advanced stages of endometriosis the impairment of NK cell activity occurs with increased E2, and decreased PRL plasma levels. Additional studies are required to determine whether the E2/PRL ratio represents a possible biochemical marker of endometriosis.

The aim of the study was to investigate the prognostic significance of estimating tumor cell proliferation in stage I cervical squamous carcinoma by analyzing MIB 1 immunostaining with respect to the lesion size, lymphatic spread, and clinical outcome. A possible relationship between MIB 1 index and natural killer activity was also discussed. The medical records of 34 patients with stage I squamous cervical carcinoma who had undergone primary radical surgery at the Institute of Gynecologic and Obstetrics, Ancona University, between 1988 and 1993, were recruited from our series of 57 consecutive cases and reviewed. Thirty-one patients were considered eligible for the study and evaluated for age, demographic characteristics, tumor histologic grade, tumor size, lymphatic spread, and adjuvant radiotherapy. The expression of primary tumor proliferation related to Ki67 antigen was immunohistochemically evaluated by monoclonal MIB 1 antibody (Immunotech, Marseille Cedex, France) on microwave oven-processed formalin-fixed paraffin-embedded tissue. The basal natural killer cell activity of peripheral blood lymphocytes was evaluated against K562 cell line and expressed in lytic units for each patient. The MIB 1 immunostaining was significantly related with tumor size (P = 0.001) and lymphatic spread (P = 0.009); in contrast, there was no relationship between grade of histologic differentiation and MIB 1 immunostaining. The Cox proportional hazards analysis showed a significant independent relationship between MIB 1 immunostaining and disease-free survival (P = 0.004). The analysis of natural cytotoxicity defined a significant inverse relationship between peripheral blood lymphocyte's natural killer activity and tumor MIB 1 immunostaining (r = -0.07, with P = 0.03). Our data defined the prognostic significance of tumor cell proliferation immunostaining, an interesting parameter correlated with the disease-free survival in locally advanced cervical carcinoma. The relationship between MIB 1 index and natural killer activity is interesting; natural cytotoxicity seems to be altered in the host with respect to the cervical carcinoma characteristics.

Our objective was to investigate the role of estrogens in the development and progression of endometriosis, and evaluate the in vitro boosting effect of lymphokines on the activity of natural killer cells from endometriosis patients, with respect to the estradiol concentrations. Natural killer activity of peripheral blood was evaluated in 42 endometriosis patients who underwent laparoscopy for pelvic pain, infertility and benign adnexal masses, and it was correlated with serum estradiol levels. Twenty-five women with moderate and severe disease were re-evaluated for immune and endocrine parameters 4-8 weeks after surgery, before any specific adjuvant medical treatment, and analyzed for in vitro responsiveness of cytotoxic cells to interferon (IFN) alpha 2 beta and interleukin-2 (IL-2) incubation. Patients with moderate and severe endometriosis showed a significant decrease of natural cytotoxicity when compared with patients with mild and minimal disease (p = 0.01). The decrease of immune reactivity was independent of a reduced representation of natural killer cells, and persisted after surgical removal of all macroscopic endometriosis foci. A significant inverse relationship was observed between natural killer activity and serum estradiol levels, which resulted in moderate and severe disease (r = -0.4, p = 0.009) but not in stages I and II. The in vitro responsiveness of cytotoxic cells to lymphokine incubation was preserved; both IFN alpha 2 beta and IL-2 were able to increase the cytotoxicity of natural killer cells significantly from advanced-stage patients (p = 0.014 and p = 0.006 for IFN alpha 2 beta and IL-2 respectively).(ABSTRACT TRUNCATED AT 250 WORDS)

Objective: To analyze longitudinally the basal natural killer cell activity in patients with invasive cervical carcinoma, the natural cytotoxicity was related to the most important known prognostic factors, and evaluated with respect to the clinical outcome of cervical disease., Materials and Methods: Forty-six patients with histologically proven invasive cervical carcinoma treated and followed at the Institute of Gynecology and Obstetrics, Ancona University, Salesi Hospital, were consecutively recruited from 1989 to 1992 and included into the study. For immunologic investigation, natural killer cell activity and peripheral blood T-lymphocyte subsets were tested before primary treatment and during the follow-up period, every 6 months. Natural killer activity was determined by target cell retention of the fluorescent dye carboxyfluorescein diacetate; the K 562 cell line was used as target cells., Results: A significant inverse relationship was observed between natural killer activity and disease stage (p = 0.001); patients with stage IV disease had the lowest level of natural cytotoxicity. The reduction of natural cytotoxicity was not accompanied by any alteration of lymphocyte distribution. The longitudinal analysis showed an increase of natural killer activity after surgical removal of the tumor, persisting during the follow-up, but all the 9 patients who recurred showed, at the time of disease recurrence, a significant decrease of their natural cytotoxic potential., Conclusion: Natural killer cell activity seems to be a functional index of immune status, significantly related to the stage and the clinical outcome of disease.

The objective was to examine the prevalence of human papillomavirus (HPV) DNA infection in mild cervical dysplasia and to evaluate longitudinally the persistence of HPV DNA positivity in an observational study, aiming at identifying the role of peripheral blood lymphocyte natural killer activity in the natural history of dysplastic disease. Twenty-three patients with histologically proven mild cervical dysplasia were selected. The HPV DNA positivity, determined by polymerase chain reaction, and cervical dysplasia were monitored cytologically and colposcopically at the 3rd (time 1), 6th (time 2) and 12th months (time 3), and defined by biopsies for routine histology taken at times 2 and 3. For each patient included in the study, the immune reactivity was evaluated at the time of diagnosis and afterwards, longitudinally during the follow-up. The immune status analysis included T lymphocyte subsets (CD3, CD4, CD8, CD56, CD16 monoclonal antibodies by Beckton Dickinson, Mountain View, Calif., USA) and determinations of natural killer cell activity (against the sensitive cell line K 562). Eighteen out of the 23 women with mild cervical dysplasia (78.3%) were found positive for HPV DNA, with a significantly high representation of HPV DNA type 16 (55.6% of cases). At the end of the study, 12 out of 18 HPV-DNA-positive women became negative (defined by two or more negative tests) for the original HPV DNA type, with 66.7% of spontaneous HPV DNA negativization rate (p = 0.6).(ABSTRACT TRUNCATED AT 250 WORDS)

It has been clearly demonstrated that immune responses may be conditioned in a manner similar to that of the classical Pavlovian experiments. Evidence of impaired immune function in aging has raised the question of whether psychological conditioning of an immune response can also be effective in old age. The knowledge that aged mice have decreased spleen cell natural killer (NK) activity and that NK cytotoxicity, at least in young mice, can be psychologically conditioned led us to explore in old mice the possibility of conditioning the response of NK cell activity using the odor of camphor as the conditioned stimulus (CS) and the injection of Poly I:C as the unconditioned stimulus (US). Young and old male mice were divided into five and six groups, respectively. They received the CS and/or the US in association (conditioning) trials (sessions 1-9). Mice were exposed to the camphor odor alone at 72 hours after the final association trial to observe the conditioning phenomenon (session 10). The group conditioned with Poly I:C and camphor and receiving the CS at session 10 showed statistically significant increases in spleen cell NK activity over those of the control groups that did not receive the CS treatment at session 10 (2.6- and 4.0-fold increase in young and old, respectively). Treatment with camphor odor alone had no effect on boosting NK cell activity. These findings demonstrate the possibility of conditioning immune responses in old age, offering a valuable tool for attenuating age-related immune deterioration in various species, including the human. In addition, these results again confirm highly significant immune enhancement by classical conditioning and extend previous findings from female mice to males as well.

The purpose of this study was to examine the relationship between natural killer cell activity and biological behavior of tumor, expressed by architectural (FIGO) and nuclear grading, depth of myometrial invasion, and proliferating cell nuclear antigen (PCNA) index in patients with endometrial carcinoma. Forty patients with FIGO stage I endometrial carcinoma, treated with radical surgery, were included in this retrospective study. At the time of diagnosis, natural killer cell activity of peripheral blood was evaluated against K562 target tumor cells and correlated with architectural and nuclear grading, depth of myometrial invasion, and PCNA index of the tumor. Natural killer activity diminished with increasing nuclear grade of the tumor (P = 0.004); similarly, natural cytotoxicity decreased with myometrial invasion: for stages IC and IB endometrial carcinoma, the mean values of natural cytotoxicity were significantly lower than stage IA disease (P = 0.0001). Natural killer activity was significantly correlated with PCNA immunostaining of the tumor (r = -0.8). It is concluded that the natural immune reactivity seems to be related to the pathologic features of early stage endometrial carcinoma, showing a significant reduction in presence of nuclear pleomorphism and/or myometrial invasion, and also an inverse relationship with PCNA index.

Objective: The aim of our study was to evaluate the basal immune reactivity in patients with locally advanced endometrial carcinoma, and the immune modulating effect of adjuvant treatment in cancer population randomized to endocrine or radiation therapy., Material and Methods: Forty-three patients with FIGO stage I and II endometrial carcinoma, treated with primary radical surgery, were randomly selected to receive endocrine (medroxy-progesterone acetate 30 mg weekly and Tamoxifen 300 mg weekly, given consecutively) or radiation adjuvant treatment (external beam photon treatment of the whole pelvis, with an average total dose of 3680 cGy rad). The immune assay included the evaluation of natural killer cell activity by target cell retention of the fluorescent dye carboxyfluoresceyn diacetate, using sensitive cell line K 562. The immunological monitoring was performed before surgery, and then before, during, and after adjuvant treatment. Nine patients, who refused any adjuvant treatment, were recruited as controls., Results: Patients, matched for age and demographic characteristics, with locally advanced endometrial carcinoma had significantly lower mean values of natural killer activity than in healthy controls; the decrease of natural cytotoxicity was significantly related to the depth of myometrial invasion. The adjuvant treatment was associated with a significant immune modulation: patients in endocrine therapeutic regimen showed an increase of natural killer activity, while patients in radiation therapy had a reduction; in the control group there was no significant modification of natural cytotoxicity during negative follow up., Conclusions: Natural killer cell activity of peripheral blood is significantly reduced in local advanced endometrial carcinoma patients. The natural cytotoxicity evaluation, as a function of therapeutic modality, may be useful in establishing a relationship between adjuvant treatment and immune status in endometrial carcinoma.

Peripheral blood natural killer (NK) cell activity against K562 tumor line was monitored in 16 patients with locally advanced squamous cervical carcinoma (II and III FIGO stages) during neoadjuvant polychemotherapy (cisplatin at 80 mg/m2 and bleomycin at 30 mg/m2). There was a significant progressive decrease in NK activity during the three cycles of antiblastic treatment (P = 0.008) without significant depletion of NK cell (CD56 and CD16 monoclonal antibody positive cell percentage and absolute number). A significant relationship was shown between basal NK activity levels and response to polychemotherapy; in fact, nonresponder patients had a significantly lower mean value of NK activity before polychemotherapy than responders (P = 0.044). In conclusion, NK activity declines as a result of antiblastic therapy; although polychemotherapy reduces tumor spread, its antineoplastic action may be affected by this decreased immune reactivity.

The objective was to evaluate natural cytotoxicity of peripheral blood during interferon treatment in cervical intraepithelial neoplasia grade 2 (CIN2), as index of interferon activity. Twenty-one patients with CIN2, histologically proven, were treated with interferon alpha 2b (Intron A, Sheering-Plough Corp.), in a dose of 3,000,000 units three times per week for 8 weeks self-administered by intramuscular injection. The rate of remission to the interferon treatment was 38.1%. Eight healthy patients had a significant increase in natural killer activity during and after the treatment (p < 0.001), whereas the 13 nonresponder patients remained with low values of natural killer activity. By analyzing the basal natural killer activity before the treatment, the patients with clinicopathologic remission had a significantly higher mean value than nonresponder patients (p = 0.007). Notwithstanding the specimen exiguity, the individual basal natural killer activity seems to be a predictive parameter of interferon treatment response in patients with CIN2.

Objective: To evaluate the correlation between natural killer cell activity and serum estradiol (E2) levels in patients with different stages of endometriosis., Methods: Natural killer cell activity of peripheral blood lymphocytes and serum E2 levels were evaluated in 73 women who underwent laparoscopy for pelvic pain, infertility, and benign adnexal masses., Results: The 33 patients (45%) with endometriosis showed a significant decrease in natural killer cell activity in relationship to an increase in disease stage (correlation coefficient r = -0.83, P < .001). A significant inverse relationship was observed between cytotoxicity and serum E2 levels (correlation coefficient r = -0.89, P < .001)., Conclusion: The relationship between natural killer cell activity and serum E2 levels suggests that an immunoendocrine interaction plays an important role in the progression of endometriosis.

A longitudinal evaluation of the natural killer (NK) cell activity of peripheral blood lymphocytes was performed in 17 patients with advanced ovarian cancer (treated surgically and with subsequent multiagent chemotherapy) for a median follow-up period of 32 months. At the time of primary treatment, the mean value of NK cell activity was significantly lower in patients who then had disease progression (p < 0.05) than in patients with progression-free survival. During the follow-up period, no significant modifications of the NK cell activity were observed; only at the time of clinical disease progression did the NK cell activity show a significant reduction. We conclude that the NK cell activity is a potentially important prognostic factor.

In the present study we investigated the effect of thyrotropin (TSH) on both the proliferative capacity and the natural killer (NK) cell activity of murine spleen lymphocytes. It was found that TSH at various concentrations significantly increased the proliferative response of mouse lymphocytes to both concanavalin A (Con A) and phytohemagglutinin (PHA). This increase was particularly evident when suboptimal concentrations of mitogens were used (40-50% increase). The administration to cell cultures of TSH alone could not induce a significant stimulation of proliferative capacity. In order to provide a better knowledge about the mechanism by which TSH improved the mitogen-induced lymphocyte proliferation, the effect of the pituitary hormone on lymphocytes directly stimulated with recombinant interleukin-2 (RIL-2), was examined. It was observed that there was a great increase in IL-2-induced lymphocyte proliferation by TSH. The improvement in proliferative capacity of lymphocytes was particularly evident by using suboptimal rIL-2 concentrations (25-30% increase). The studies carried out on the cytotoxic activity of NK cells showed that TSH was able to significantly increase the IL-2-induced NK cell activity without modifying the basal levels of cytotoxicity. The results support the immunoregulatory role of TSH and contribute towards understanding the mechanisms of interaction between neuroendocrine and immune systems.

We have studied the systemic T-lymphocyte subset and, in particular, the natural-killer (NK) activity in 23 patients with cervical intra-epithelial neoplasia (CIN), and 11 controls (a history of normal cervical PAP smears and colposcopy patterns). The lymphocyte phenotypes were studied using a panel of monoclonal antibodies, and NK activity was evaluated as the percentage lysis of K 562 target cells. In patients with CIN we found a significant decrease in NK activity (p = 0.002) which was directly correlated with the grade of CIN, with no alterations in the absolute number of NK cells.

Basal natural killer (NK) cell activity in peripheral blood lymphocytes (PBLs) of 29 patients with gynecologic malignancy and 10 healthy controls was performed using K 562 cell line. A significant decrease of NK cell activity was observed in patients with gynecologic malignancy (p = 0.003). The NK cell activity of patients with poorly differentiated tumor or with stage IV of disease was significantly reduced with respect to patients at any other lower grade or earlier stage. We conclude that in patients with gynecologic malignancies there is a decrease of basal NK cell activity of PBLs, which is more significant in cases of distant tumor dissemination or poorly differentiated tumor (p less than 0.001).

Old mice show a reduced natural killer (NK) cell activity. Among the causes proposed to explain this defect, both intrinsic failure of NK cells or age-related alteration of microenvironmental factors relevant for NK function, have been taken into consideration. The findings reported in the present paper, demonstrate that thymic peptides, and in particular the facteur timique serique (FTS), more recently called thymulin (ZnFTS) in its zinc-bound form, whose production and activity is generally reduced in old age, is able, when administered in vitro, to restore the crippled NK cytotoxicity of spleen cells from old mice. Neither the zinc-unbound form of the hormone (FTS) nor zinc ions alone are effective. The action is exerted on the basal NK activity and not on IFN-boosted NK cytotoxicity, at variance with the restoration obtained in similar experimental conditions with thyroid hormones, thus suggesting that different mechanisms or maturative steps of NK cells are involved.

The cytotoxic activity of Natural Killer (NK) and Lymphokine Activated Killer (LAK) cells in neoplastic patients with or without antalgic treatment was studied. NK cell activity was found reduced in untreated neoplastic patients when compared to healthy subjects. The atalgic treatment with morphine (orally or intrathecally administered) was able to significantly reduce the mean values of NK cell activity found in cancer patients. In three patients the cytotoxicity of NK cells significantly decreased during transfer from oral to intrathecal administration of morphine. In contrast to the NK cell function, the development of LAK cell activity significantly increased in neoplastic patients when compared to healthy controls. Further increments were obtained during treatment with morphine. The oral treatment with morphine was able to determine a higher induction of LAK cells than the intrathecal administration of the drug. Besides providing new knowledge on the effect of morphine on immune system our findings suggest that, in order to include neoplastic patients in clinical trials of adoptive immunotherapy with LAK cells and interleukin-2 (IL-2), the antalgic therapy with oral administration of morphine may represent a better solution than the intrathecal administration of the drug.

The age-dependent changes in thyroid-hormone blood levels and the effects of in vivo and in vitro thyroid-hormone administration on both basal and lymphokine-induced spleen cell natural killer (NK) activities have been investigated in young and old Balb/c mice. Both thyroxine (T4) and triiodothyronine (T3) plasma levels decline progressively with increasing age of the mice, displaying in 25-month-old mice only 50 and 60% of the T4 and T3 blood levels, respectively, found in young mice. In vivo T4 administration to old mice causes a significant increment in endogenous NK activity (2.2-fold increase), which approaches the values observed in young animals, while it does not modify NK activity in young mice. The T4 injection in old mice does not induce changes in the lymphocyte sub-populations. When T4 is administered in vitro alone or in combination with interferon (IFN) and/or interleukin 2 (IL-2), no effect is observed either on basal activity or IL-2-induced cytotoxicity, whereas the IFN sensitivity of spleen cells from old mice is significantly recovered (4-fold increase). T4 is able to increase IFN-induced cytotoxicity even when administered in vitro simultaneously with IFN to the cytotoxic assay (1.5- and 2.7-fold increases in young and old mice, respectively). Under these conditions, IFN alone is not able to exert any boosting effect even at a young age. In vivo propylthiouracil (PTU) administration completely abrogates the IFN responsiveness of spleen cells in young mice. The interruption of the PTU treatment results in a recovery of IFN-inducible NK cytotoxicity. Taken together, our findings point out the important role of thyroid hormones in the modulation of NK cell activity and provide a new insight into the mechanisms by which the endocrine system is able to influence the expression of natural immunity.

Several abnormalities arise at the level of NK cell function in both rodents and humans with increasing age. The age-related changes are not irreversible alterations since they can be reversed either by endocrinological or nutritional approaches, suggesting that age-related microenvironmental changes may play a relevant role in the age-related immune deterioration. Whether endocrine and nutritional factors have an additive effect or act through the same intracellular mechanisms remains to be established, though the first possibility seems more likely since the action of TSH and thyroid hormones is specifically directed towards lymphokine-boosted NK activity, while AL are able to prevent age-associated defect of basal NK cytotoxicity.

Results of several experiments have given rise to the hypothesis that the decline of the immunocompetence with aging is at least in part related to alterations of the lipid membrane composition and, consequently, to a decrease in membrane fluidity. The age-dependent decline of mitogen responsiveness can, in fact, be reversed by a special lipid mixture designated as active lipids (AL 721), which acts by means of its fluidizing action on the plasma membrane. The purpose of this study was to examine the possibility of raising the low endogenous levels of Natural Killer (NK) activity by in vitro AL administration in old mice. When spleen cells from old mice were incubated in vitro with AL, a significant increase in cytotoxic activity was obtained over control cultures, without reaching, however, the levels observed in young mice. In spleen cells from young mice, the AL administration causes a slight augment of NK basal activity. These results suggest that cell membrane fluidity plays an important role in the efficiency of NK cells, giving support to the hypothesis that a rectification of rigidified cell membranes may represent a valuable approach to restore proper physiological functions in old age.

The possibility of increasing the responsiveness of lymphoid cells to Interferon (IFN) was investigated by studying the effect of the in vitro preincubation of spleen cells from young mice with thyroid hormone prior to the administration of IFN on their Natural Killer (NK) activity. Both physiological concentrations of T4 or T3 are able to significantly increase the boosting action of IFN on NK cells. The increase in IFN-responsiveness is observable at various IFN concentrations and the peak level of IFN-induced NK activity is reached with less than a half-optimal IFN concentration after T4 pretreatment. The effect of T4 on the IFN-sensitivity of NK cells requires a very short time, being already fully expressed after only 30 min, with minor increments at more prolonged incubation times. These results indicate for the first time the possibility of modulating the in vitro sensitivity to IFN by a hormonal approach.

Of the immunological alterations in pregnancy hypertension were studied peripheral blood lymphocyte subpopulation in normal and hypertensive pregnancies by means of monoclonal antibodies. In pregnant women with hypertension an increase was found in NK activity. It was also shown that an increase in circulating T cells expressing Tac antigen occurred in women with pregnancy hypertension. These preliminary data on Tac antigen suggest that there is an activation of Leu 3 cells; which may introduce the concept of Leu 3 activity like NK activity. Further studies on this subject could explain that the concept of Leu 3 activity is correct.

In the present study we observed the absolute number of NK cells and the NK activity in 10 patients with cervical cancer. NK activity was determined before and after stimulation with Active Lipids. We reported a significant decrease of basal NK activity, but after AL stimulation NK activity was increased with statistical significance. Therefore our findings show that the reduction of NK activity in neoplastic patients is not an irreversible phenomenon since it can be restored by "in vitro" AL administration. This is important for the adoptive immunotherapy by which we could give LAK cells to the patients.

The influence of "in vitro" treatment with thyroia hormones on basal or lymphokine-induced NK cytotoxicity was analyzed in Balb/c mice using YAC-1 as target cells in a 51 Cr release assay. "In vitro" treatment with IFN or IL-2 causes a considerable increase of the natural cytotoxic activity of spleen cells from young-middle aged mice. An additive effect is observed when the two lymphokines are administered simultaneously. "In vitro" spleen cells preincubation with thyroxine increases NK citotoxic activity induced by IFN but it does not modify the basal activity or that induced by IL-2. Furthermore, thyroxine was able to significantly amplify even the maximal boosting effect induced by the simultaneous addition of both IFN and IL-2. These findings suggest a role for thyroid hormones in the modulation of natural cell-mediated citotoxicity and particularly in the espression of NK cell sensitivity to IFN.

The age-dependent modifications of both basal and lymphokine-induced natural killer (NK) activity were analyzed in Balb/c inbred mice by measuring either the percentage of specific lytic activity in a 51 Cr release assay or the percentage of cells capable of binding the YAC-1 target cells. The basal lytic activity of spleen cells is low at birth, then it increases progressively and reaches a peak between 5 and 8 weeks of age and decreases thereafter, displaying in 25-month-old mice only 10% of the NK activity found in young mice. The number of spleen cells capable of binding target cells is higher at birth and then it declines progressively-old mice show about 40% of the binding capacity found in young mice. Significant in vitro NK activation is obtained in spleen cells from 25-, 50- and 750-day-old mice by incubation with interleukin-2 (IL-2) (1.8-fold increase in 25- and 50-day-old mice and 2.6-fold increase in 750-day-old mice) while no effect is obtained in 15-day-old mice. The responsiveness to in vitro stimulation with interferon (IFN) is not present in 15-day-old mice, however it appears in a defective way at the age of 25 days, and reaches its highest level in 50- and 180-day-old mice, (3.5- and 4.5-fold increase), still remaining present in 630-day-old mice, (two-fold increase) but not in 750-day-old mice. The in vivo injection with IFN increases the spleen cell NK activity of 25- and 50-day-old mice (3.6- and 2.3-fold increase respectively) but not of 15- and 750-day-old mice. The present data here confirm the existence of age-dependent variations of spleen cell NK activity and suggest a similar sequential timing of appearance/disappearance of IL-2 and IFN responsive spleen cells during ontogenetic development and aging respectively.