Matrix metalloproteinase profile in patients with Creuztfeldt-Jakob disease.

Preliminary findings suggest that abnormalities in matrix metalloproteinase (MMP) activity may be found in the cerebrospinal fluid (CSF) of patients with Creutzfeldt-Jakob disease (CJD). In this study of 16 subjects with CJD and 16 age-, and sex-matched controls, we determined the presence of MMP-2 and MMP-9 in their active and proenzyme forms, the relative levels of MMP-3 and four inhibitors of MMP activity (TIMP-1, TIMP-2, TIMP-3 and TIMP-4), and the concentration of 4-3-3 protein. The methodology used involved zymography and immunological techniques. The results indicate that, compared with controls, CJD patients have a significantly higher positive frequency of pro-MMP-9 and of the active form of MMP-2, along with significantly higher levels of TIMP-1 and TIMP-2, classical inhibitors of MMP-9 and MMP-2, respectively. We also found a positive correlation between 14-3-3 protein concentration and that of TIMP-1 and TIMP-2 levels (correlation coefficients of 0.793 and 0.798, respectively). These results suggest that abnormalities in MMP and TIMP profiles may be helpful in the biochemical characterisation of CJD.

Effect of Salmonella-infected human monocytes on natural killer cell cytotoxicity. In vitro studies.

Various chemicals, including some bacteria-derived components, modulate natural killer cell (NKC) activity. We have analyzed the effect of wild-type Ty2 and of mutant strain TYT1231 Salmonella typhi-infected monocytes (U937 cells and human autologous monocytes) on NKC cytotoxicity of peripheral blood mononuclear cell (PBMC) and highly purified NKC (HPNKC; CD16+/56+ > 95%; the rest corresponding to CD3+ T-cells). PBMC's co-culture with either S. typhi strain infected U937 cells (medium or non-infected U937 cells as controls) resulted in the induction of lymphocyte activated killer (LAK) cell activity showing cytotoxicity against target human NKC-resistant lymphoblastoid Daudi cell line. Comparable experiments using autologous monocytes gave similar results. Co-culture of HPNKC preparations with either S. typhi strain infected U937 cells resulted in increased LAK cell activity against target Daudi cells in each and everyone of the five samples tested; paired Student's t-test p < 0.01 for both times (20 and 40 h) tested. Similar to the results observed in the experiments using PBMC, we did not find significant differences in the ability between medium and non-infected cells, or between wild-type S. typhi Ty2 and mutant strain TYT1231 infected U937 cells, to induce LAK activity in HPNKC preparations. PBMC co-incubation with either S. typhi strain infected U937 cells or autologous monocytes resulted in significant increases in IL-12, TNF-alpha, and IFN-gamma secretion. In similar experiments using HPNKC samples instead, infected U937 cells significantly increased IL-12 and IFN-gamma, but not TNF-alpha secretion. PBMC co-incubation with non-infected U937 cells, but not with non-infected monocytes, significantly increased supernatant IL-12 and TNF-alpha levels (no significant changes in IFN-gamma were recorded). Secreted cytokines remained essentially unchanged after co-incubating HPNKC preparation with non-infected U-937 cells. Incubation of PBMC or HPNKC preparations with either S. typhi strain infected U937 cells failed to produce significant changes in the expression of NKC lineage (CD16+/56+) or activation (CD28+, CD69+ and CD95+) markers. The ability of infected monocytes to induce LAK activity, release NKC cytokines and upmodulate NKC's CD95+ marker expression was essentially the same for both infecting Salmonella strains used. These results suggest a role for NKC in the physiological defensive response against intracellularly infected monocytes representing, perhaps one of the earliest antimicrobial mechanisms of the innate immune system.

Hypercalcemia, arrhythmia, and mood stabilizers.

Recent findings in a bipolar patient receiving maintenance lithium therapy who developed hypercalcemia and severe bradyarrhythmia prompted the authors to conduct a retrospective study of bipolar patients with lithium-associated hypercalcemia. A printout of all cases of hypercalcemia that presented during a 1-year period was generated. After eliminating spurious hypercalcemias or those associated with intravenous fluids, the authors identified 18 non-lithium-treated patients with hypercalcemias related to malignancies and other medical conditions (group A) and 12 patients with lithium-associated hypercalcemia (group B). Patients in group B were not comparable to those in group A, as the latter were medically compromised and were receiving multiple pharmacotherapies. Thus, two control groups were generated: group C1, which included age- and sex-comparable lithium-treated bipolar normocalcemic patients, and group C2, which included bipolar normocalcemic patients treated with anticonvulsant mood stabilizers. The electrocardiographic (ECG) findings for patients in group B were compared with those of patients in groups C1 and C2. It was found that these groups did not differ in their overall frequency of ECG abnormalities; however, there were significant differences in the frequency of conduction defects. Patients with hypercalcemia resulting from medical diseases and bipolar patients with lithium-associated hypercalcemia had significantly higher frequencies of conduction defects. Patients in group A had significant mortality at 2-year follow-up (28%), in contrast to zero mortality in the other three groups. The clinical implications of these findings are discussed.

Effect of Salmonella typhi wild type and O-antigen mutants on human natural killer cell activity.

We investigated the effect of glutaraldehyde-fixed Salmonella typhi Ty2 (Vi(-)) wild-type (World Health Organization's vaccine strain) and mutant strains MEI028 (rough, O-antigen(-)) and MEI012 [smooth (O-antigen(+)95%), immunomagnetically isolated NK cell preparations. Incubation of PBMC with each and every one of the S. typhi strains studied consistently and significantly, increased this cellular immune function, as well as the supernatant level of the various cytokines tested e.g. IFN-gamma, TNF-alpha, IL-10 and IL-12 (ELISA). In similar experiments, a significant increase in the cytolytic activity of HPNK cells was elicited by S. typhi Ty2 but not by mutant strain MEI028; neither of the cytokines assayed (IFN-gamma and TNF-alpha) was detected in the supernatant. Our results suggest that S. typhi O-antigen plays an essential role in a mechanism resulting in the direct activation of NK cell activity in HPNK cell preparations. However, the relative quantitative significance of this antigen in the direct stimulation of NK cell cytotoxicity expression in PBMC samples is less clear, as it appears that in this case bacterial-induced monocyte-released cytokines plays a most important role. Incubation with S. typhi Ty2 or MEI028 elicited significant expression of CD69, an early marker of NK cell activation, in PBMC but not in HPNK cell samples (flow cytometry); in similar experiments, the expression of CD16/56 and activation marker CD25 remained essentially unchanged.

Degradation kinetics of leucine5-enkephalin by plasma samples from healthy controls and various patient populations: in vitro drug effects.

Incubation of [3H]tyrosine leucine5-enkephalin with platelet-poor human plasma (final concentration 1 x 10(-8) M; 1:9 ratio to Trizma base buffer, pH 7.4) resulted in rapid and complete peptide degradation in each of the subjects studied, with more than 95% of the initial labeled tyrosine consistently recovered as the free amino acid (< or =30 minutes). Essentially, and irrespective of the incubation time (1-180 minutes), tyrosine was the only Leu metabolite formed; we were unable to identify significant amounts (> or =3%) of any other possible labeled or nonlabeled Leu degradation fragments. Neither gender (64 men and 20 women; samples tested individually), age (men, 23-70; women, 25-65 years), nor the subjects' medical condition appeared to make a significant difference in either the t1/2 of Leu elimination, the initial velocity of this reaction (x +/- SD, median, minimum and maximum of 12.0 +/- 0.9, 12.0, and 10.6-13.7 minutes; 1.2 +/- 0.3, 1.1, and 0.6-2.0 pg/min, respectively), or in the Km and Vmax values for aminopeptidase Leu degradation (x +/- SD; 0.81 +/- 0.01 mM and 14.30 +/- 1.17 micromol/L/min, respectively). Subjects were diagnosed as chronic schizophrenics (n = 15), polydrug abusers including alcohol (n = 9) and polydrug abusers excluding alcohol (n = 8), chronic alcoholics (n = 12), and migraineurs (n = 10) during or outside an acute migraine episode; for comparison we used a group of gender-matched (20 men and 10 women), age-comparable, drug-free, healthy volunteers. Differences in plasma storage time or repeated sample freezing and thawing failed to alter significantly any of these kinetic parameters of Leu metabolism or to change the identity and/or relative ratio of the products formed. The Leu degradation rate was pH and temperature dependent (optimum, 7.4 and 37 degrees C, respectively). Leu degradation was strongly and similarly inhibited by puromycin, bacitracin, and bestatin (IC50 [+/- SD] of 1.4 +/- 0.2 micromol/L) and to a lesser extent by various L-tyrosine-containing Leu fragments. The kinetics of this reaction was not significantly affected by either thiorphan, N-carboxyphenylmethyl leucine, or any other of a number of monoamine neurotransmitters, substances of abuse, nonsteroidal anti-inflammatory agents, and miscellaneous compounds tested (concentration up to 10(-4) mol/L).

Gelatinase activity of matrix metalloproteinases in the cerebrospinal fluid of various patient populations.

We have studied the enzymatic gelatinolytic activity of matrix metalloproteinases (MMPs) present in cerebrospinal fluid (CSF) of samples obtained from 67 individuals, twenty-one nonneurological patients (considered controls) and 46 subjects with various neurological disorders e.g., vascular lesions, demyelination, inflammatory, degenerative and prion diseases. Biochemical characterization of MMPs, a family of neutral proteolytic enzymes involved in extracellular matrix modeling, included determination of substrate specificity and Ca+2 dependency, as well as the effects of protease inactivators, carboxylic and His (histidine) residue modifiers, and antibiotics. Whereas all CSF samples expressed MMP-2 (gelatinase A) activity, it corresponded in most cases (normal and pathological samples) to its latent form (proenzyme; pMMP-2). In general, inflammatory neurological diseases (especially meningitis and neurocisticercosis) were associated with the presence of a second enzyme, MMP-9 (or gelatinase B). Whereas MMP-9 was found in the CSF of every tropical spastic paraparesis patient studied, its presence in samples from individuals with vascular lesions was uncommon. Patients blood-brain barrier damage was ascertained by determining total CSF protein content using both, the conventional polyacrylamide gel electrophoresis procedure under denaturing conditions and capillary zone electrophoresis.

Flow cytometric analysis of lymphocyte subsets in migraine patients during and outside of an acute headache attack.

We have conducted flow cytometric studies of two subsets of lymphocyte markers in groups of migraineurs during (n = 12; group B) and outside (n = 10; group C) of a migraine without aura attack (total n = 22; group A), including a group of patients tested in both of these phases (n = 5; group D), and compared these results with those obtained from a population of age-comparable, sex- and race-matched healthy volunteers (n = 12; group E). Comparison of the first set of lymphocytes (CD3+CD16 + 56+, CD3-CD16 + 56+, CD3-CD19+, CD3+CD19+, and CD3+HLA - DR+) between the patients in group A and the controls (group E) showed differences, reflecting greater group A percentages of CD3+CD16 + CD56+ and CD3-CD19+ lymphocytes. Furthermore, these differences reached statistical significance only for the CD3+CD16 + CD56+ lymphocytes, and then solely for the patients in group C (Scheffe's test, p < 0.05). Paired analysis of the above lymphocyte markers for subjects in group D failed to show significant differences between patients when they were having and not having a migraine attack, raising the possibility that results from a larger study could show meaningful increases in percentages of CD3+CD16 + CD56+ lymphocytes as one of the immune parameters useful for differentiating migraineurs from controls. Comparison of a second set of lymphocyte markers (CD19+CD5+, CD20+CD72-, CD20-CD72+, CD20+CD72+) among either the different groups of patients or between the patients and controls failed, however, to show statistically significant differences, emphasizing the apparent specificity of the findings described above for CD3+CD16 + CD56+ lymphocytes. Our results, albeit of a preliminary nature, suggest the occurrence of significant, differential changes in lymphocyte subset immunophenotyping between groups of pain-free migraineurs and patients during an acute migraine episode or controls. Corroboration of these findings may prove useful in clinical laboratory practice to identify changes in immunological parameters specifically associated with migraineurs, and help towards a better understanding of the etiology and pathophysiology of this condition.

In vitro effect of bacterial lipopolysaccharide on the cytotoxicity of human natural killer cells.

Preincubation with a number of mediators of infection, such as Gram negative bacteria (S. typhi), bacterial lipopolysaccharide (LPS), tumor necrotic factor-alpha (TNF-alpha), and interleukin-2 (IL-2), significantly increases natural killer (NK) cell activity in samples of human peripheral blood mononuclear cells (PBMC), without changing the levels of either the phenotypic CD16/56 or stimulatory CD25 marker. We now report similar results after preincubation of highly purified NK cell preparations (CD16 + 56 > 95%; the rest corresponding to CD3+ T-cells) with either S. typhi, TNF-alpha or IL-2. However, in similar experiments, LPS inhibits, in a dose-dependent manner (final conc. 2.5, 5.0 or 10.0 micrograms/mL), NK cell cytotoxicity against K-562 tumor cells. Preincubation of purified NK cells with LPS (25 micrograms/mL; 10 and 30 min) produced significant alterations in the tyrosine phosphorylation/dephosphorylation pattern of several intracellular proteins, including a significant increase (10 min) in the phosphorylation of the 120; 100; 72 and 59 kDa proteins, followed (30 min) by the essentially complete desphosphorylation of the p59 protein. Qualitatively similar results were obtained at lower LPS concentrations e.g., range 2.5 to 20 micrograms/mL. The absence of phosphoproteins in the 40-44 kDa range, known to be present after incubation of monocytes with LPS, raises the possibility that these "class" of proteins may be critical in explaining the LPS inhibitory effect on NK lytic function. Our finding may contribute to a better understanding of the mechanisms involved in the complex in vivo interaction between LPS, monocytes and NK cells.

Effects of the administration of amphetamine, either alone or in combination with reserpine or cocaine, on regional brain beta-phenylethylamine and dopamine release.

The effect of amphetamine sulfate (AMPH) on beta-phenylethylamine (PEA) and 3-methoxytyramine (3MT) levels in the rat frontal and cingulate cortices, the nucleus accumbens, and the striatum were evaluated after the administration of either cocaine or reserpine alone and in combination with AMPH. The purpose of this study was to evaluate the neuromodulator properties of PEA on dopamine (DA) release as reflected by 3MT steady-state concentrations. The highest concentration of PEA was found in the nucleus accumbens, followed by the cingulate and frontal cortices, and then the striatum. Time-course effects of the intraperitoneal administration of 5 mg/kg AMPH on PEA and 3MT concentrations were similar but not identical. AMPH at a dosage of 1 mg/kg significantly increased PEA concentration only in the striatum. A dosage of 2.5 mg/kg reserpine, which markedly depressed 3MT levels in all brain regions studied except the striatum, significantly reduced PEA concentrations only in the nucleus accumbens. This dosage of reserpine reduced DA concentrations by more than 80% in all regions examined, but its effects on norepinephrine were less marked. Pretreatment with cocaine (10 mg/kg) or reserpine (2.5 mg/kg) potentiated the effects of 1 mg/kg AMPH on PEA and 3MT levels in the frontal cortex and of 3MT in the striatum. Pretreatment with either 1 mg/kg reserpine (specifically used to partially mobilize DA storage) or cocaine (10 mg/kg) produced quantitative changes in the effects of 5 mg/kg AMPH on PEA and 3MT levels that were region-specific. For example, in contrast to the cortical regions and the nucleus accumbens, the AMPH-induced increase in 3MT was potentiated in the striatum. On the other hand, the increase in brain PEA produced by AMPH (5 mg/kg) was not influenced by either increased cytoplasmic DA (as deduced from the effects of 1 mg/kg reserpine pretreatment) or DA uptake inhibition (as deduced from the effect of cocaine pretreatment) in the frontal cortex or the nucleus accumbens. Furthermore, the increase in PEA produced by AMPH (5 mg/kg) in the cingulate cortex and the striatum were abolished and potentiated, respectively, by these drug pretreatments. Our results suggest that although DA release and PEA formation are stimulated by AMPH, these effects appear to involve mechanisms that are not directly related and hence suggest a dissociation between 3MT and PEA formation in the brain. Our work also suggests that PEA is most likely not to be co-released with DA following the administration of AMPH. Therefore, it is concluded that whatever physiological role PEA may play in central synaptic transmission, its effects do not appear to be dependent on DA release.

Enhancement of natural killer cell activity in HIV-1-infected subjects by a mixture of the calcium ionophore A23187 and the phorbol ester TPA: lack of response to a similar challenge with interleukin-2 or alpha-interferon.

When compared to controls (n = 30), human immunodeficiency virus type-1 (HIV-1)-positive individuals, either asymptomatic (n = 10) or diagnosed with acquired immunodeficiency syndrome (AIDS) (n = 10), showed a statistically significant decrease in the percentage and absolute number of CD4 ( + ) T-lymphocyte cells (flow cytometry, Becton Dickinson FACScan; mean +/- SD of 42.6 +/- 6.9 and 948.5 +/- 393.3, 19.5 +/- 8.7 and 269.8 +/- 174.3, 4.6 +/- 4.1 and 60.1 +/- 134.3, respectively; Student's t- test, p < 0.05). However, this decrease was less marked in asymptomatic patients; in fact, the percentage and number of the above cells in this group of subjects was significantly higher than in the AIDS patients (Student's t-test, p < 0.05). However, we failed to find significant differences in the percentage of natural killer cells (NKCs; CD15 ( + ) CD56 ( + ) ) between the HIV-1-infected asymptomatic or AIDS groups of patients, or when compared with the controls (mean +/- SD of 10.4% +/- 9.4%, 14.3% +/- 9.7%, and 14.8% +/- 6.4%, respectively). Whereas either group of patients had a lower number of NKCs per microliter than the control group (mean +/- SD of 137.8 +/- 87.6, 91.1 +/- 98.3, and 331. 5 +/- 266.5, respectively), this decrease only reached statistical significance for the AIDS patients (Student's t-test, p < 0.05). Healthy controls showed statistically significantly higher NKC activity than either the HIV-1-infected asymptomatic or AIDS group of patients (K-562 target cell; mean +/- SD and range values as percentage of specific lysis of 19.1% +/- 15.6% and 2.4%-58.2%, 3.4% +/- 3.2% and less than 0.1% [non-detectable]-10.3%, and 6.4% +/- 5. 5% and less than 0.1%-19.5%, respectively; Student's t-test, p < 0. 05). Challenge of samples from the control group with either interleukin-2, alpha-interferon, or with a mixture of the calcium ionophore A23187 (Io) plus the 12-O-tetradecanoylphorbol-13-acetate ester (TPA) resulted in every case in a statistically significant increase in NKC lytic function (mean +/- SD and range values as percentage of specific lysis of 19.1% +/- 15.6% and 2.4%-58.2%, 27. 6% +/- 17.4% and less than 0.1%-56.0%, 32.1% +/- 20.9% and 2.1%-76. 4%, and 62.6% +/- 24.0% and 16.7%-95.0%, respectively; Student's t-test, p < 0.05). A similar challenge for samples from the HIV-1-positive subjects, either asymptomatic or with AIDS, resulted in most cases in an enhanced NKC activity; however, this increase in NKC lytic function reached statistical significance only for the group of Io + TPA-incubated samples (Student's t-test, p < 0.05). These results indicate that control or patient baseline NKC activity, and the response of this cellular immune function to a challenge with different immunomodulators, are phenotype-independent. They also suggest an association between HIV-1 infection and alterations in the initial mechanisms responsible for NKC activation; a similar general explanation has been suggested to account for the abnormal NKC lytic function observed in various severe pathological conditions, e.g., extensive burns, polytrauma, and sepsis. Understanding the molecular mechanism involved in regulating initial NKC activation could provide the rational basis for the design of newer pharmacological strategies to treat these conditions.

Activation of protein tyrosine kinase: a possible requirement for fixed-bacteria and lipopolysaccharide-induced increase in human natural killer cell activity.

Preincubation of peripheral blood lymphocytes (PBL) from drug-free, healthy volunteers with either the protein tyrosine kinase inhibitor genistein (GNT, n = 10, final concentration 200 microM) or the protein kinase A activator dybutiryl-cyclic-AMP (cAMP, n = 11, final concentration 10 microM), resulted in a significant inhibition of natural killer cell activity (NKCA, expressed as percentage of specific chromium release). With the exception of 4 out of the 11 cAMP-treated samples, individual values for NKCA in the drug preincubated specimens were at least 20% below the same subject baseline activity; furthermore, NKC lytic function was non-detectable in 4 out of the 10 and in 1 out of the 11 samples pretreated with either GNT or cAMP, respectively. PBL preincubation with glutaraldehyde-fixed Gram-negative bacteria (GNB, n = 13, final GNB-to-effector cell ratio of 50 : 1) resulted in a statistically significant increase in NKCA (baseline (x +/- SD) of 21.6 +/- 16.4 and bacteria treated samples of 41.5 +/- 24.6, respectively, Student's paired t-test p < 0.05). At least a 20% increase in NKC lytic function over its own baseline value was recorded for 11 out of the 13 samples tested (Table 1). Preincubation with GNB and GNT (5 samples) not only blocked the immunostimulant effects of GNB (Student's paired t-test p < 0.05), but in most cases individual values for NKCA were similar to those recorded for GNT-only treated samples. Use of cAMP instead of GNT also blocked, but to a smaller extent, the GNB-produced increases in NKC lytic function (paired Student's t-test < 0.05). PBL preincubation with lipopolysaccharide (LPS, n = 11, final concentration 50 micrograms/ml) resulted in a statistically significant increase in NKCA (baseline (x +/- SD) of 20.7 +/- 14.1 and LPS treated samples of 39.2 +/- 18.5, respectively, Student's paired t-test < 0.05). At least a 20% increase in NKCA over its own baseline value was observed for each and everyone of the 11 samples studied (Table 2). Addition of LPS and GNT to the incubation mixture resulted not only in inhibition of the NKCA upmodulating LPS effects (Student's paired t-test p < 0.05), but each and everyone of the individual samples' NKCA were, in fact, significantly lower than their corresponding control baseline values and similar to those recorded for GNT-only treated samples. However, the use of LPS and cAMP (Table 2) produced less dramatic results, significant inhibition of LPS effect were recorded in only 2 samples (Nos 8 and 10), and individual NKCA in the remaining 3 specimens was significantly higher than the corresponding baseline value. Whereas experimental results obtained with GNT support the involvement of PTK-dependent pathways in the stimulation of human NKCA produced by GNB and LPS, cAMP experiments suggest modulation of PKA-dependent pathways as responsible for the decrease in NK lytic function produced by a number of chemicals involved in the pathophysiology associated with certain forms of stress, including septic shock. Further research in this area could help in the rational design of pharmacological approaches for the treatment of these conditions.

Studies of natural killer cell activity in a drug-free, healthy population. Response to a challenge with taxol, estramustine and lipopolysaccharide.

Preincubation of peripheral blood lymphocytes with the microtubule disturbing agents estramustine (ETMN; n = 7, final conc. 20 microM) or taxol (TX; n = 13, final conc. 10 microM), resulted in a statistically significant inhibition of natural killer cell activity [(NKCA); baseline (x +/- SD; expressed as percentage of specific chromium release) of 32.2 +/- 30.5 and 34.4 +/- 27.7 and drug treated samples of 13.9 +/- 19.9 and 12.5 +/- 20.8, respectively; Student's paired t-test p < 0.005]. Furthermore, most individual values for NKCA in the drug preincubated samples were at least 20% below the same subject baseline lytic function (except for TX sample No.1), and NKCA was non detectable in 4 out of 7 and 5 out of 13 samples (pretreatment with either ETMN or TX< respectively). The use of other concentrations and different preincubation times for these chemotherapeutic agents also produced NKCA inhibition, which was time and dose dependent. Preincubation with lipopolysaccharide (LPS; n = 16, final conc. 50 micrograms/ml), an endotoxin prominently involved in the etiology of septic shock, resulted in a statistically significant enhancement of NKCA [baseline (x +/- SD; expressed as percentage of specific chromium release) of 25.4 +/- 20.4 and LPS treated sample of 36.6 +/- 17.4, respectively; Student's t paired t-test p < 0.005]. At least a 20% increase in NKC lytic function over its own baseline value was recorded for each and everyone of the samples tested with LPS.2+ the pathophysiology associated with septic shock.

Improvement of axial dystonia with the administration of clozapine.

A 45-year-old white male with a positive family history for various neurological disorders (not including dystonias), and a long history of neuroleptic exposure on account of a chronic schizoaffective illness, developed severe torsion trunkal movement to the left which were accompanied by inversion of the left arm and outward extension of the right arm, as well as left torticollis. The axial dystonia, which was refractory to treatment, was disabling interfering with the activities of daily living, posture and gait. Radiological studies revealed marked dextroscoliosis. The administration of clozapine in dosages of 200 mg p.o. in the morning and 250 mg p.o. at bedtime, resulted in a significant improvement of the neck and trunk dystonia. After the discontinuation of the clozapine, an exacerbation of the movement disorder back to baseline levels prior to the use of this agent, was observed. Subsequent therapy with thioridazine in a dosage of 600 mg/day did not mask or treat the dystonia. The clinical implications of these findings are discussed.

Enhancement of natural killer cell activity in septic shock patients by a mixture of the calcium ionophore A23187 and the phorbol ester TPA: in vitro studies.

Preincubation of peripheral blood lymphocytes from drug-free, healthy volunteers with a mixture of the calcium ionophore A23187 (Io) and the phorbol ester TPA (12-O-tetradecanoylphorbol-13-acetate) consistently resulted in a significant enhancement (dose-dependent; maximum immunostimulation obtained with the Io + TPA final mixture concentration of 10 uM + 250 ng/ml, respectively) of natural killer cell activity (NKCA) (n = 8; mean +/- SD of 16.8 +/- 8.9 and 52.0 +/- 18.0, paired Student's t-test p < 0.005; effector-to-target cell ratio of 30:1). Results from the same protocol, but using samples from septic shock patients followed a similar trend; however, and perhaps reflecting the significantly lower baseline NKCA in this group of individuals (n = 7), the mean value reached for this cellular immune function after incubation with Io + TPA was significantly lower than that of the treated controls' group (mean +/- SD of 19.8 +/- 11.6 and 52.0 +/- 18.0, respectively, Student's t-test p < 0.005). As expected from the role of calcium in the activation of NKCA, incubation with the Io significantly increased baseline NKCA, which was largely unchanged by TPA. Expression of the CD56+ and CD16+ phenotypes in septic shock patients did not correlate directly with NKCA, suggesting that this condition may be associated with changes in the function rather than the quantity of these cellular markers.(ABSTRACT TRUNCATED AT 250 WORDS)

Cutaneous and ocular changes associated with the use of chlorpromazine.

A 57-year-old schizoaffective disorder patient was placed on chlorpromazine because of its sedative properties and low profile for extrapyramidal side effects. After two years of treatment, the patient developed photosensitivity and blue-gray pigmentation of the skin of the face, neck and dorsum of hands. Significant pigment deposits were also observed in eye lens and cornea. An oval translucent lesion lateral to the left angle of the mouth was removed and the biopsy showed a basal cell carcinoma. The cutaneous and eye pigmentation were only partially reversible after the discontinuation of chlorpromazine. There has been no recurrence of basal cell carcinoma.

In vitro studies of natural killer cell activity in septic shock patients. Response to a challenge with alpha-interferon and interleukin-2.

Natural killer cell activity (NKCA) in patients with septic shock was statistically significantly lower than the value recorded for a group of drug-free, healthy volunteers [9.1 +/- 7.8 (n = 20) and 20.6 +/- 16.6 (n = 15), respectively; Student's test, p < 0.05]. As expected, preincubation of peripheral blood lymphocytes from samples taken from a group of controls with either alpha-interferon or interleukin -2 resulted in an enhancement of NKCA for each and everyone of the subjects studied; however, results from a similar protocol using patient samples showed a lack of consistency, both in the direction and magnitude, in the elicited changes in NK lytic function. Whereas samples from same patient responded with either an increase or a decrease in NKCA to preincubation with both immunostimulators, others responded with NKCA upmodulation to one and downmodulation to other of these test substances. A better knowledge of the mechanism(s) responsible for the depressed expression of NKCA in septic shock patients, and its altered response to alpha-interferon and interleukin-2, could generate new modalities in the diagnosis and therapy of this condition.

In vitro studies of natural killer cell activity in post traumatic stress disorder patients. Response to methionine-enkephalin challenge.

Attempts to define biological parameters that may be specifically associated with the pathophysiology of post traumatic stress disorder (PTSD) have met with only limited success, reflecting perhaps the practical limitations resulting from the high frequency of comorbidity of this condition with Axis I, II and III psychiatric diagnoses. We now report our studies on natural killer cell activity (NKCA), and the response of this cellular immune function to an in vitro methionine-enkephalin (MET) challenge in a population of Vietnam veterans with PTSD. Due to the characteristics of our PTSD patients, our protocol included four sex-matched, age-comparable control groups: (1) chronic alcoholics, (2) chronic drug abusers excluding alcohol, (3) chronic users of alcohol and other drugs of abuse and (4) drug-free, healthy volunteers. Although these groups did not significantly differ in their "baseline" NKCA, significant findings emerged from their response to preincubation with MET (10(-10), 10(-8) and 10(-6) M; 40:1 effector-to-target cell ratio). To minimize interindividual variations in the expression of NKCA each subject was used as its own control. Whereas peptide challenge resulted in an increase in NK lytic function in a subpopulation of group four (one or more MET concentrations, 8 out of 22 subjects), it produced mixed results in samples from individuals in group 2, and in general failed to elicit NKCA changes in the samples from participants in groups 1 and 3. Nine of the thirteen PTSD patients responded to MET preincubation with decreases in NKCA, which in five of them reached values below 20% of baseline for the three peptide concentrations tested. These findings may suggest that the "stress factor" in Vietnam veterans with PTSD plays a role in downmodulating NK lytic function in response to an in vitro MET challenge, an effect that appears to be potentiated by the use of drugs of abuse other than alcohol. The possible clinical relevance of these findings, including the identification of a subgroup of PTSD patients on the basis of immunological tests such as the one described in this work, deserves further investigation.

Enhancement of human natural killer cell activity by opioid peptides: similar response to methionine-enkephalin and beta-endorphin.

We studied the effect of methionine-enkephalin (MET) and beta-endorphin upon human peripheral blood lymphocyte natural killer cell (NKC) activity in a group of healthy volunteers (n = 27; 17 male and 10 female, age +/- SD and range of 32 +/- 6, 25-43 years and 36 +/- 11, 22-65 years, respectively). Aliquots from some individual samples were preincubated separately with different concentrations of either peptide (n = 12), while others were tested with only one of these substances (MET, n = 6; beta-endorphin, n = 9). Using each individual as its own control, MET (10(-8) and 10(-6) M) and beta-endorphin (10(-10) and 10(-8) M) significantly increased NKC activity (NKCA) (at least 20% over base value, effector-to-target cell ratio, 40:1) in 7 out of 15 and 7 out of 19 subjects, respectively. Results obtained from the rest of the samples were mixed, e.g., changes observed in NKCA were not significant or showed significance with only one of the peptide concentrations studied. Cells from individuals showing a significant increase in NKC lytic function following preincubation with either MET or beta-endorphin responded similarly to the other peptide (in both cases 5 out of 6 subjects), suggesting that enhancement of NKCA by MET and beta-endorphin may work through a similar mechanism.