Body composition and -174G/C interleukin-6 promoter gene polymorphism: association with progression of insulin resistance in normal weight obese syndrome.

Insulin resistance and obesity are intimately related to a chronic low grade systemic inflammation. Interleukin-6 (IL-6) may influence the pathogenesis of obesity-related diseases. The aim of this study is to investigate the effect of body's fat mass on the relationships between -174G/C IL-6 promoter gene polymorphism, IL-6 circulating level and insulin resistance. A population of 150 Caucasian women was studied, subdivided according to their body composition in non-obese (NW), Normal Weight Obese (NWO) and preobese-obese (OB). The NWO subjects were found in an intermediate position between the NW and OB subjects in terms of body weight, fat mass percentage (FM%), abdominal FAT%, hs-CRP and plasma triglyceride level. Fasting plasma IL-6 concentration was positively correlated with the homeostasis model assessment for insulin resistance (HOMA-IR) in all subjects analyzed (P=0.0014). In NWO and OB women a significantly increased IL-6 mean value was observed compared with NW subjects. In G/G population, the IL-6 plasma level of NWO and OB was significantly higher with respect to NW. No significant differences of IL-6 concentrations were observed in the three groups carrying G/C genotype. NWO and OB women homozygous for the allele C have significantly lower value of IL-6 with respect to NW subjects. IL-6 concentration was positively correlated with FM% in G/G (R(2)=0.397, P<0.001) and was negatively correlated in C/C (R(2)=0.459, P=0.002). No significant correlation was observed in G/C genotype (R(2)=0.041, P=0.173). In conclusion our study confirms that, at least in Italian Caucasian females, the FM% is a major determinant of an increase in IL-6 production and insulin resistance. -174 G/C IL-6 promoter polymorphism represents a marker which could help to identify, time in advance, "vulnerable" individuals at risk of age and obesity related diseases.

Normal Weight Obese syndrome: role of single nucleotide polymorphism of IL-1 5Ralpha and MTHFR 677C-->T genes in the relationship between body composition and resting metabolic rate.

We have identified a subset of metabolically obese, but normal weight individuals, with potentially increased risks of developing the metabolic syndrome, despite their normal body mass index. We determined the relationship among body fat distribution, resting metabolic rate (RMR), total body water amount (%TBW), selected gene polymorphism on interleukin-15 receptor-alpha (IL-15Ralpha) and methylenetetrahydrofolate reductase 677C-->T (MTHFR 677C-->T), to distinguish normal weight obese (NWO) from nonobese with a normal metabolic profile and obese individuals. We analysed anthropometric variables, body composition by Dual energy X-ray Absorptiometry (DXA), RMR by indirect calorimetry, %TBW by bioimpedence analysis (BIA), MTHFR 677C-->T and IL-15Ralpha genotypes of 128 clinically healthy Caucasian individuals. We compared a group of female, defined as NWO and characterised by a BMI < or = 25 kg/m(2) and FM > or = 30% with groups of others female, and males, represented by nonobese with a BMI < or = 25 kg/m(2) and FM < or = 30%, and preobese-obese individuals with BMI > or = 25 kg/m(2) and %FM > or = 30%; none of the males was classified as NWO. Significant correlations were found among body fat mass distribution, metabolic variables, percentage of total body water distribution and selected genetic variations. The variables that contributed significantly to the separation of classes were body tissue (Tissue), %TBW, RMR, the volumes of both oxygen (VO2) and carbon dioxide (VCO2). The distribution of MTHFR 677C-->T and IL-15 genotypes was significantly different between classes. Our data highlight that NWO individuals showed a significant relationship between the decrease in the basal metabolism (RMR), body fat mass increasing and total water amount. Possession of wild type homozygotes genotypes regarding IL-15Ralpha cytokine and 677C-->T MTHFR enzyme characterised NWO individuals.

Body composition analyses in normal weight obese women.

The purpose of this study was to identify new indexes of body composition that characterize the normal weight obese (NWO) women. We measured body composition by dual energy x-ray absorptiometry (DXA) and resting metabolic rate (RMR) by indirect calorimetry in a cohort of seventy-five healthy Italian women, subdivided into three groups (nonobese/controls, NWO, preobese-obese women). Despite a normal body mass index (BMI), the NWO women have a higher body fat mass percentage (FAT %) (38.99 +/- 6.03) associated to a significant (p = 0.02) lower amount of lean mass of legs (12.24 +/- 1.31) and lean mass of left leg (6.07 +/- 0.64) with respect to the control group. The NWO group showed a significant (p = 0.043) lower RMR (1201.25 +/- 349.02) in comparison with nonobese and preobese-obese women. To classify NWO individuals among general population, we identified three significant body composition indexes: abdominal index, leg index and trunk index. The NWO women showed significant increased value in the three indexes (p < 0.001). Our results suggest that, despite a normal BMI, the NWO women displayed a cluster of anthropometric characteristics (body fat mass percentage, leg indexes) not different to obese women ones. An appropriate diet-therapy and physical activity may be protecting NWO individuals from diabetes and cardiovascular diseases associated to preobese-obese women.

The immunosuppressive cytokines influence the fetal survival in patients with pregnancy-induced hypertension.

PROBLEM: The present study examines the hypothesis that the elevated levels of transforming growth factor (TGF)-beta1 and interleukin (IL)-10 would be protective for the fetus survival during pregnancy-induced hypertension (PIH). Moreover, we evaluate the IL-12 and IL-15 serum concentrations and their relationships with PIH. METHOD OF STUDY: Serum samples were obtained before the onset of labor from control and PIH groups. Cytokine concentrations were determined by Enzyme-Linked Immunoadsorbent Assay. RESULTS: Our data show that PIH women have significantly higher TGF-beta1 and IL-10 concentrations with respect to control groups (P = 0.0001). Similarly, macrophages from the PIH placentas produce in vitro more elevated TGF-beta1 and IL-10 levels compared to normal pregnant ones (P = 0.02), also in the absence of LPS stimulation. IL-12 and IL-15 serum concentrations were not detectable in all pregnant groups. CONCLUSION: We have found that PIH women have elevated concentrations of anti-inflammatory/immunosuppressive cytokines, suggesting their important role in fetal allograft protection during the normal and pathological pregnancy.

9-(2-Phosphonylmethoxyethyl) adenine increases the survival of influenza virus-infected mice by an enhancement of the immune system.

PMEA (9-(2-phosphonylmethoxyethyl)adenine) is a potent inhibitor of DNA viruses and retroviruses able to enhance natural immune functions such as natural killer cell activity and interferon production. The results reported in this paper show that the treatment with PMEA significatively decreased the mortality of mice challenged with influenza A/PR8 virus (an RNA virus, non sensitive to the antiviral effect of PMEA) compared to untreated, infected controls (median survival 8.64 days and 7.61 days, respectively), and reduced lung weight and consolidation (two surrogate markers of virus infection). Furthermore, virus titer obtained from lung homogenates was substantially decreased in PMEA-treated mice compared to controls. Finally, enhancement of natural killer cell activity was achieved in PMEA-treated A/PR8-infected mice compared to A/PR8-infected controls. Overall, results suggest that PMEA decreases the influenza virus-related mortality and morbidity through the enhancement of some immune functions, and that this effect might be additive or even synergystic with the direct inhibitory effect of DNA viruses or retroviruses induced by PMEA itself. This supports the importance of evaluating this drug in patients with diseases related to herpesviruses or to human immunodeficiency virus.

Enhancement of natural killer activity and interferon induction by different acyclic nucleoside phosphonates.

Acyclic nucleoside phosphonate (ANP) analogues are a class of compounds with potent activity against herpesviruses and/or retroviruses. Our preliminary experiments have shown that 9-(2-phosphonylmethoxyethyl)adenine (PMEA), a prototype of the ANP family, enhances some parameters of natural immunity. In this paper we have evaluated the effect of different schedules of administration of PMEA and other ANP analogues of clinical interest upon natural killer (NK) activity and interferon (IFN) production in a mouse model. The results show that PMEA significantly enhances NK activity and interferon production. Other ANP analogues tested in our system, i.e., 9-(2-phosphonylmethoxyethyl)-2,6-diaminopurine (PMEDAP), and 9-(3-fluoro-2-phosphonylmethoxypropyl)adenine (FPMPA), similarly induced enhancement of natural immunity. The immunomodulating effect of PMEA was even more pronounced with a single administration compared to repeated administrations of the drug. Dose-dependent enhancement of NK activity and IFN production could also be demonstrated during chronic administration of PMEA (more resembling to what will be the schedule of administration of this drug in patients). Overall, the data here presented suggest that the enhancement of some natural immune functions induced by ANP analogues may add to the direct antiviral activity of these drugs against retroviruses and herpesviruses, and thus may be able to increase the host resistance against viral infections.

Immunomodulatory activity of 9-(2-phosphonylmethoxyethyl)adenine (PMEA), a potent anti-HIV nucleotide analogue, on in vivo murine models.

In order to evaluate the influence of antiviral nucleoside analogues upon the natural immune system, we investigated the immunomodulatory activity of 9-(2-phosphonylmethoxyethyl)adenine (PMEA), a nucleotide analogue with potent anti-HIV and anti-herpes activity, in a murine system. C57BL/6 mice were inoculated intraperitoneally with 10, 25 and 50 mg PMEA/kg. Mononuclear cells were isolated from their spleens, and some natural immune functions were evaluated. The results show that PMEA significantly increases the levels of natural killer (NK)-cell cytotoxicity. We also found that alpha/beta IFN production was substantially increased in PMEA-treated mice, while both IL-1 and IL-2 production was decreased. Thus, PMEA can increase some natural immunity functions, such as NK activity and IFN production. These results suggest that PMEA might be active in vivo against HIV and herpes viruses both as an immunomodulator and as an antiviral compound.

Suppressor cells induced by influenza virus inhibit interleukin-2 production in mice.

PR8 virus depressed interleukin-2 (IL-2) and natural killer (NK) cell activity in BALB/c infected mice. IL-2 production was not dependent on (i) a decreased number of T cells or (ii) a primary defect in IL-1 production, but on a T-suppressor cell subpopulation. In fact, when T suppressor cells were removed from infected spleen cells, we observed normal levels of IL-2 activity.

Pinealectomy inhibits interleukin-2 production and natural killer activity in mice.

Four--five-week-old C57BL/6 mice were surgically pinealectomized. At different time intervals after surgery their spleens were removed and assayed for interleukin-2 (IL-2) production and natural killer (NK) cell activity. Non-operated and sham-operated mice were used as controls. The present results indicate that pinealectomy significantly reduced IL-2 production and NK cell activity, in comparison to sham-operated mice. These effects seem to be related to the lack of melatonin. In fact the subcutaneous injection of this hormone (50 or 100 mg/kg at 5 p.m.) in pinealectomized mice was able to restore IL-2 production and NK cell activity. However, chronic treatment with melatonin (10, 20 and 50 mg/kg for 9 consecutive days) failed to reverse the impairment of the immune responses.

Surface glycoproteins of influenza virus increase cell-mediated immunity functions in mice.

T-lymphocyte blastogenic response to Concanavalin A, interleukin-2 (IL-2) production and NK cell activity were studied in three groups of Balb/c mice infected with intranasal inoculum of A/Chile/1/83 (H1N1) influenza virus, vaccinated with same virus glycoproteins and infected fifteen day after vaccination. Virus infection results in T-cell function impairment. In fact T-cell blastogenic response and IL-2 production are profoundly decreased as early as 24 hours, whereas NK cell activity is depressed only later. Instead viral glycoprotein vaccination induces an enhancement of CMI related parameters and protects the host from virus immunosuppressive effects.

PR8 influenza virus infection impairs serum thymic activity levels and thymus-derived immune functions in mice.

The levels of serum thymic activity (STA), the Thy-1.2 positivity of spleen "spontaneous" rosette-forming cells (SSRFCs) (measured in terms of Az sensitivity), as well as the blastogenic response to specific mitogens for T-lymphocytes, were studied in Balb/C mice after intranasal infection with A/PR/8/34 (HON1) influenza virus. As early as 12 hours, and more drastically, 24 hours the levels of STA were profoundly decreased after virus infection. Spleen Az sensitivity and blastogenic response of thymocytes and splenocytes to stimulation with Concanavalin A and Phytohemagglutinin, respectively, were depressed only later (day 2 or 3). These changes remain evident for about 1 week and later revert to normal values. All of the effects described are dose-dependent and appear to be virus related. Thence the PR8 virus infection initially induces a decrease of STA levels and secondly a impairment of thymus-derived immune functions.

Immunofluorescence study of thymuses of mice given Friend leukemia virus: conventional vs monoclonal antibodies.

In the course of a study of the early effects of Friend Leukemia Virus (FLV) infection in thymus structure and function, evidence of early localization of infectious FLV in the thymic type I and type II epithelio-reticular cells of susceptible mice was obtained. Such evidence was based upon bio-assay, ultrastructural and immunofluorescence observations. As for the latter, conventional monospecific sera against FLV p30 and gp70 antigens as well as two distinct monoclonal antibodies recognizing FLV gp70 epitopes were employed. Both monoclonal antibodies stained with a granular pattern the cytoplasm of type I and II epithelio-reticular cells from susceptible mice injected with live FLV. On the contrary, conventional monospecific sera diffusely stained the cytoplasm of all epithelio-reticular cells of the thymus, independently of mice inoculation with and susceptibility to virus, possibly recognizing tissue-associated normal mouse antigens and/or cross-reacting antigens of other ecotropic viruses.

[Effects of thymosin alpha l on some time-dependent functions in thymectomized mice]. / Effetti della timosina alfa 1 su alcune funzioni timo-dipendenti in topi timectomizzati.

The effects of a synthetic thymosin alpha1 on the azathioprine-sensitivity of spleen cells and on the thymic-like activity of serum from adult thymectomized mice, were observed. Thymosin alpha1 is able to restore the lowered levels of these T-dependent functions after in vivo administration.

Effects of in vivo Friend leukemia virus infection on levels of serum thymic factors and on selected T-cell functions in mice.

The levels of serum thymic factor(s) (STF), of Thy-1.2 positivity of splenocytes [as measured by their azathioprine (AZ) sensitivity], and of Thy-1.2-positive "spontaneous" spleen rosette-forming cells (SSRFCs), as well as the presence of infectious virus in the thymus, were assessed as a function of time after virus inoculation in susceptible DBA/2, partially resistant BALB/c, and fully resistant C57BL/6 mice given the polycythemia- or anemia-inducing strain of Friend leukemia virus (FLV-P and FLV-A, respectively). As early as Days 2 to 3, the levels of STF and of AZ sensitivity of splenocytes were profoundly decreased in DBA/2 mice, and, to a lesser extent, in BALB/c mice given FLV-P; however, SSRFCs/spleen were increased in both mouse strains. Conclusive evidence of infectious FLV-P was obtained in the thymuses of DBA/2 mice soon after infection. In mice of the same strains infected with FLV-A, STF levels were similarly decreased, but AZ sensitivity of splenocytes was unaffected, and SSRFCs were decreased. Evidence of early FLV-A infection in the thymus of DBA/2 mice was likewise obtained. In C57BL/6 mice given FLV-A, STF levels, AZ sensitivity of splenocytes, and SSRFC showed changes similar to, but of lower magnitude than, those in BALB/c mice. On the other hand, in C57BL/6 mice given FLV-P, the decrease in STF and AZ sensitivity was almost as pronounced as in susceptible DBA/2 mice in the face of complete absence of infectious virus or viral markers in the thymuses. The observed changes are ascribed to virus infection in view of the following: (a) good temporal correlation between these changes and virus infection; (b) absence of any change in mice given heat-inactivated viruses or spleen homogenate of normal DBA/2 mouse spleen; (c) overall good correlation between mouse genotype and genetic (Fv-1 and Fv-2) restrictions of virus infection on one hand and the magnitude of the observed changes on the other. In particular, the decrease in STF and SSRFC levels is ascribed to the replication-competent (Friend-murine leukemia virus) component of Friend leukemia virus complex, whereas the decrease in AZ sensitivity of splenocytes and the increase of SSRFCs are ascribed to the defective spleen focus-forming virus component of the complex. All changes described so far were transient, since they were not detectable beyond 42 days after virus inoculation in overtly leukemic animals. The observed derangements of thymus-derived immune functions may play an important cofactor role during the onset of leukemia in mice genetically permissive to Friend leukemia virus replication and transformation, but they do not seem relevant to the maintenance of leukemia.

Modulation of endogenous prostaglandins by thymosin-alpha 1 in lymphocytes.

The effects of thymosin-alpha 1 on the stimulation of specific release of prostaglandin E2 (PGE2) from splenic lymphocytes and thymocytes were studied. Experiments were also performed to study in parallel the absolute levels of thymosin-alpha 1 in the blood and the induction of serum FTS activity and of azathioprine sensitivity of spleen cells from adult thymectomized (ATx) mice. A significant difference in the release of PGE2 between normal splenocytes and splenocytes from ATx mice was observed. Thymosin-alpha 1 at certain concentrations was able to stimulate PGE2 release from lymphocytes of ATx mice while inhibiting release in lymphocytes of normal mice. Also, thymocytes were stimulated to release PGE2 after incubation with alpha 1 in a manner similar to that seen in spleen cells of ATx mice. Approximately the same concentrations of alpha 1 was found to also correct the low azathioprine sensitivity of splenocytes from ATx mice. Determinations of FTS-like activity in the blood and the pharmacokinetics of alpha 1 after administration of this synthetic molecule show a clear dissociation. A maximum peak of alpha 1 activity was obtained after 1 hr, while maximal FTS-like activity was observed after 24 hr. The inhibition of the induction by alpha 1 of FTS-like activity and of Thy 1.2 antigen by indomethacin suggests that the action of alpha 1 requires prostaglandin biosynthesis.

Is thymosin action mediated by prostaglandin release?

Treatment of spleen cells derived from adult thymectomized mice with thymosin fraction 5 resulted in a rapid and dose-dependent stimulation of the release of immunoreactive prostaglandin E2. The release of prostaglandin E2 was associated with induction of theta antigen and was totally inhibited by indomethacin. In contrast, prostaglandin E2 release from spleen cells from intact donors was inhibited by treatment with fraction 5. The data support the concept that prostaglandin E2 mediates the effects of thymosin fraction 5 on lymphocytes.