Prophylaxis and therapy of HBV infection in 20 patients treated with disease modifying antirheumatic drugs or with biological agents for rheumatic diseases.

OBJECT: To evaluate the safety and tolerability of lamivudine in patients with HBV infection needing immunosuppressive treatment for rheumatic diseases. PATIENTS AND METHODS: Twenty patients with rheumatic diseases planned to receive immunosuppressive DMARDs or biological agents were screened for HBV markers. In all active carriers antiviral treatment was recommended. Inactive carriers (HBsAg positive, aminotrasferase and viremia persistently normal) were divided into two risk categories according to the type and the degree of immunosuppression, and antiviral prophylaxis was started only in patients of the high risk category. Antiviral treatment was recommended also in potential occult carriers (HBsAg negative, HBcAb positive) treated with rituximab. In twenty patients antiviral treatment was started: 1 was a potential occult carrier planned to receive rituximab; 9 were inactive carriers, in which prophylactic therapy was needed for a high risk of HBV reactivation (in 3, for the use of TNF blocking agents); 10 were treated for active viral replication. Prophylaxis and therapy were performed with lamivudine. In three patients adefovir was associated. RESULTS: Antiviral drugs were well tolerated. In all cases, immunosuppressive treatment was given for the planned duration of therapy, with good results on the rheumatic diseases. Median duration of antiviral treatment was 19 months (for a total of 386 month/person). No cases of viral reactivation were observed. CONCLUSION: Our experience demonstrates the feasibility of a prophylaxis and therapy of HBV infection in patients with rheumatic diseases. This approach reduces the risk of viral reactivation and allows the choice of the optimal immunosuppressive treatment in rheumatic patients.

[Early rheumatoid arthritis: a prospective study on how to induce the remission]. / Induzione della remissione delle poliartriti reumatoidi all'esordio: studio prospettico.

OBJECTIVES: To investigate whether the aggressive use of DMARDs can control the clinical disease in the early arthritis, to define new parameters of the disease aggressivity and to study the effectiveness of RMN in comparison with RX focusing on the articular erosions. METHODS: 45 patients having a case of early arthritis (less 6 months) with 3 or more swollen joints were recruited and treated with 80 mg of steroids in order to distinguish persistent arthritis from non persistent ones. Afterward we began to use DMARDs with persistent arthritis and, if it wasn't helpful, we shifted to anti-TNFalpha therapy. The clinical response was valued by SDAI. RESULTS: After 1 year our therapeutic approach showed a remission in 60% of the patients. The 82% of remaining obtained a significant SDAI improvement and only in 3 cases we used anti-TNFalpha due to a persistent high disease activity. Anti-CCp were positive in 46% of patients in remission and in 53% of the rest. The bone erosions were present in 4 patients only and they were detected by RMN, only 2 by RX. CONCLUSIONS: We observed a clinical remission in the 60% of patients treated with aggressive DMARDs. During our trial, anti-CCp weren't predictive about the therapy response. We observed that RMN is more effective than RX in detecting erosions and it's necessary for diagnosis and follow-up of early arthritis.

[Anti-TNFalpha agents in elderly patients with rheumatoid arthritis: a study of a group of 105 over sixty five years old patients]. / I farmaci biologici anti-TNFalpha nel paziente anziano affetto da artrite reumatoide: studio di una coorte di 105 pazienti ultra-sessantacinquenni.

OBJECTIVE: To evaluate the efficacy and the safety of anti-TNF alfa treatment in elderly patients (>/=65 years old) with active rheumatoid arthritis (RA), in comparison with younger (17-65 years old). METHODS: We considered retrospectively 295 patients, affected by RA and treated with anti-TNF alfa drugs. They were divided in two groups, according to their age, and followed up for two years: over-65-years old patients (190) and under-65-years old patients (105). Effectiveness of drugs was assessed analyzing RA disease activity (DAS28, DAS44, SDAI), functional status (HAQ) and serological parameters (CRP) before and after anti-TNF alfa therapy. Safety was studied considering discontinuation rate of biological disease-modifying antirheumatic drugs, and collateral events rate. RESULTS: At baseline, elderly patients showed higher disease activity's score (DAS 28, DAS44, SDAI, HAQ) with important loss of articular function (worse quality of life, HAQ) than younger patients (p<0.05). During the therapy, improvement in clinical parameters was observed (DAS28, DAS44 and SDAI) with no significant difference between the two groups. In elderly patients disability index, on the contrary, improved less than in younger (p<0.05). After treatment, also CRP decreased less in elderly patients (p<0.05). During the follow-up, 74 over-65-years old patients (38.95%) and 116 under-65-years old patients (38.05%) discontinued anti-TNF alfa therapy because of loss of efficacy (20.52% vs 11.42%), severe adverse events (17.34% vs 25.67%), voluntary discontinuation or good clinical response (1% vs 0.95%). No differences were shown about the frequency and reasons of anti-TNF alfa withdrawal (p>0.05). CONCLUSIONS: Anti-TNF alfa treatment was efficacious and safe in both groups of patients. These drugs induced improvement in disease activity, apart from the age. No functional improvement was observed in HAQ, showing the irreversible loss of articular function and the incomplete recovery in elderly patients. Age doesn't interfere with the possibility to treat elderly patients with anti-TNF alfa drugs.

Real-world experience of tocilizumab in rheumatoid arthritis: sub-analysis of data from the Italian biologics' register GISEA.

To assess the long-term effectiveness and safety of tocilizumab, abatacept, and tumor necrosis factor-α inhibitors (TNFi), in the Italian real-world setting of rheumatoid arthritis (RA). The records of adult RA patients from the Italian biologics' registry Gruppo Italiano Studio Early Arthritis (GISEA) were analyzed. Demographic and clinical data were obtained at entry. The disease remission rate (28-joint disease activity score calculated using the erythrocyte sedimentation rate [DAS28-ESR] ≤ 2.6) and frequency of adverse events (AEs) were evaluated at 2 years. From 1999 to 2014, 7539 patients were treated with biologics (61.3% in first- and 22.6% in second-line), 68% of cases received TNFi, 9.1% tocilizumab, and 8.6% abatacept. Treatment groups showed a similar DAS28 at entry. As first-line, tocilizumab induced a significantly higher remission rate than abatacept or TNFi at 6 (51 vs 23.3 and 26.2%, respectively; p < 0.0001) and 24 months (52.3 vs 33.3 and 34.4%, respectively; p < 0.01). A similar pattern was observed in later lines. The most common AEs reported were infections, reactions to biologics (more frequent among TNFi-treated patients), increased transaminase (more frequent among TCZ-treated patients), and cardiovascular events. In clinical practice, TCZ induced a rapid and long-lasting remission and in a higher percentage of patients compared to abatacept and TNFi, with a good safety profile.

Inflammatory molecules: a target for treatment of systemic autoimmune diseases.

Inflammation is a process to protect the host against infection and danger signals. However, many pathologic conditions, including autoimmune diseases, are sustained by perpetual activation of the inflammatory process. In the past few years our knowledge about the molecular basis of inflammation have been uncovered and now much is known about the primary role of pro-inflammatory cytokines such as IL-1 and TNF. In the early '90s, anti-cytokine therapies started and confirmed the primary role of TNF in autoimmune diseases, such as rheumatoid arthritis, Crohn's Disease and psoriasis. Increasing understanding of the role of inflammatory mediators in inflammation and diseases is opening new avenues for the treatment of inflammatory-based diseases through selective targeting of cytokines and lipid mediators.

Selective melanocortin MC4 receptor agonists reverse haemorrhagic shock and prevent multiple organ damage.

BACKGROUND AND PURPOSE: In circulatory shock, melanocortins have life-saving effects likely to be mediated by MC4 receptors. To gain direct insight into the role of melanocortin MC4 receptors in haemorrhagic shock, we investigated the effects of two novel selective MC4 receptor agonists. EXPERIMENTAL APPROACH: Severe haemorrhagic shock was produced in rats under general anaesthesia. Rats were then treated with either the non-selective agonist [Nle4, D-Phe7]-melanocyte-stimulating hormone (NDP--MSH) or with the selective MC4 agonists RO27-3225 and PG-931. Cardiovascular and respiratory functions were continuously monitored for 2 h; survival rate was recorded up to 24 h. Free radicals in blood were measured using electron spin resonance spectrometry; tissue damage was evaluated histologically 25 min or 24 h after treatment. KEY RESULTS: All shocked rats treated with saline died within 30-35 min. Treatment with NDP--MSH, RO27-3225 and PG-931 produced a dose-dependent (13-108 nmol kg-1 i.v.) restoration of cardiovascular and respiratory functions, and improved survival. The three melanocortin agonists also markedly reduced circulating free radicals relative to saline-treated shocked rats. All these effects were prevented by i.p. pretreatment with the selective MC4 receptor antagonist HS024. Moreover, treatment with RO27-3225 prevented morphological and immunocytochemical changes in heart, lung, liver, and kidney, at both early (25 min) and late (24 h) intervals. CONCLUSIONS AND IMPLICATIONS: Stimulation of MC4 receptors reversed haemorrhagic shock, reduced multiple organ damage and improved survival. Our findings suggest that selective MC4 receptor agonists could have a protective role against multiple organ failure following circulatory shock.

Anti-TNF-alpha treatment in rheumatoid arthritis with anti-Ro/SSA antibodies. Analysis of 17 cases among a cohort of 322 treated patients.

OBJECTIVE: To compare the efficacy and safety of anti-TNF-alpha treatment in RA patients with and without anti-Ro antibodies, in order to detect any change in their immunological or clinical profile. METHODS: Autoantibodies in 322 patients being treated with anti-TNF-alpha drugs were studied; 17 were found to be anti-Ro positive, while 305 were anti-Ro negative. RESULTS: Two groups, comparable in terms of sex distribution, RA duration and anti-TNF-alpha drug employed, showed symmetrical, erosive polyarticular RA with high disease activity. Anti-TNF-alpha led to significant improvement in both groups. At baseline rheumatoid factor and ANA, globally positive in 68.6% and 40%, were more frequent in anti-Ro positive sera. ANA showed a rising trend beginning in the 6th month of treatment in both groups, which was always statistically significant compared to baseline. Anti-dsDNA antibodies, measured using either CLIFT and ELISA or the Farr assay, remained stable in the first 6 months, then increased at 12th and 18th month, and subsequently declined. No difference was detected between the two groups regarding the number or cause of dropouts, but lupus-like disease was more frequent in anti-Ro positive subjects (p = 0.012). In addition, two cases of NHL were detected. CONCLUSION: Anti-TNF-alpha treatment was shown to be effective in patients with anti-Ro antibodies. Although anti-dsDNA and lupus-like disease were more frequent in anti-Ro positive patients, severe manifestations of systemic involvement were not observed. A longer follow-up is warranted to evaluate the risk of NHL in these patients.

[TNF-alpha inhibition in anti-Ro/SSA positive patients with rheumatoid arthritis: clinical and immunologic effects]. / Inibizione del TNFalpha in pazienti con anticorpi anti-Ro/SSA e artrite reumatoide: analisi clinica e sierologica.

OBJECTIVE: To analyse efficacy and safety of anti-TNFalpha treatment in 17 patients with rheumatoid arthritis (AR) and anti-Ro antibodies, in order to detect difference in clinical and immunological response. METHODS: 322 patients, affected by RA and treated with anti-TNFalpha drugs, were considered, searching every 6-12 months ANA, anti-dsDNA and anti-ENA antibodies. Seventeen were anti-Ro positive and 305 anti-Ro negative before starting treatment. RESULTS: Anti-Ro positive subjects showed active arthritis at baseline (mean DAS: 5), with frequent extra-articular features, such as ocular and oral sicca symptoms. They showed rapid and stable improvement during the treatment, with-out significant difference compared to anti-Ro negative group. A good clinical Eular response was shown in 46% of anti-Ro negative subjects, steady stable during time. On the contrary, fewer anti-Ro positive patients seem to be "good" responders. RA remission (DAS <1,6) was achieved in 9-25% of anti-Ro positive and 21-29% of anti-Ro negative, without significant difference. Antinuclear antibodies tend to increase in both groups, during the time. Anti-DNA increased to 40% of anti-Ro positive sera since 6th month, while they slightly increased in first 12 months in anti-Ro negative ones, then decreased to baseline value. No differences were shown about the frequency and reasons of anti-TNFalpha withdrawal, except for cutaneous lupus-like disease, more detected in anti-Ro positive group. CONCLUSIONS: Anti-TNFalpha drugs are effective in anti-Ro positive RA as well as other RA patients. Anti-DNA positivity and lupus-like disease were more frequently observed in anti-Ro positive group.

Nuclear factor-kappaB as a target of cyclosporin in acute hypovolemic hemorrhagic shock.

BACKGROUND: Cyclosporin is an immunosuppressive drug that blocks Nuclear Factor kappaB (NF-kappaB) activation. We investigated the role of NF-kappaB in acute hypovolemic hemorrhagic (Hem) shock and the effects of cyclosporin in this model of experimental shock. METHODS: Hem shock was induced in male anesthetized rats by intermittently withdrawing blood from an iliac catheter over a period of 20 min (bleeding period) until mean arterial blood pressure (MAP) fell and stabilized within the range of 20-30 mmHg. Two minutes after bleeding cessation, animals received intravenously cyclosporin (1 mg kg(-1)) or its vehicle. Survival rate and survival time were evaluated for 120 min after bleeding was discontinued. Plasma TNF-alpha levels were investigated at different time points after bleeding cessation. Moreover we investigated levels of TNF-alpha mRNA in the liver, vascular reactivity, liver NF-kappaB binding activity and levels of the inhibitory protein IkappaBalpha in the cytoplasm. RESULTS: Hemorrhagic shocked rats died in 27+/-6 min following the cessation of bleeding, experienced a marked hypotension (mean arterial blood pressure=20-30 mmHg) and had enhanced plasma levels of Tumor Necrosis Factor-alpha (208+/-22 pg ml(-1), 20 min after the end of bleeding). Furthermore, aortas taken 20 min after bleeding from hemorrhagic shocked rats showed a marked hypo-reactivity to phenylephrine (PE: 1 nM-10 microM) compared with aortas harvested from sham shocked rats. Hem shocked rats also had increased levels of TNF-alpha mRNA in the liver (15-20 min after the end of bleeding). Electrophoretic mobility shift assay showed that liver NF-kappaB binding activity increased in the nucleus 10 min after the end of hemorrhage and remained elevated until the death of animals. Western blot analysis suggested that the levels of inhibitory protein IkappaBalpha in the cytoplasm decreased at 5 min after the end of bleeding. Cyclosporin inhibited the loss of IkappaBalpha protein from the cytoplasm and prevented NF-kappaB binding activity in the nucleus. Furthermore, cyclosporin increased survival time (118+/-7 min; P<0.01) and survival rate (vehicle=0% and cyclosporin=80%, at 120 min after the end of bleeding), reverted the marked hypotension, decreased liver mRNA for TNF-alpha, reduced plasma TNF-alpha (28+/-7 pg ml(-1)), and restored to control values the hypo-reactivity to PE. CONCLUSIONS: Our results suggest that acute blood loss (50% of the estimated total blood volume over a period of 20 min) causes early activation of NF-kappaB which triggers an inflammatory cascade leading to a fatal outcome. Cyclosporin blocks NF-kappaB activation and protects against hypovolemic hemorrhagic shock.

Oxidative stress causes nuclear factor-kappaB activation in acute hypovolemic hemorrhagic shock.

Nuclear Factor kappaB (NFkappaB) is an ubiquitous rapid response transcription factor involved in inflammatory reactions and exerts its action by expressing cytokines, chemokines, and cell adhesion molecules. We investigated the role of NF-kappaB in acute hypovolemic hemorrhagic (Hem) shock. Hem shock was induced in male anesthetized rats by intermittently withdrawing blood from an iliac catheter over a period of 20 min (bleeding period) until mean arterial blood pressure (MAP) fell and stabilized within the range of 20-30 mmHg. Hemorrhagic shocked rats died in 26.3 +/- 2.1 min following the discontinuance of bleeding, experienced a marked hypotension (mean arterial blood pressure = 20-30 mmHg) and had enhanced plasma levels of Tumor Necrosis Factor-alpha (200 +/- 15 pg/ml, 20 min after the end of bleeding). Furthermore, aortas taken 20 min after bleeding from hemorrhagic shocked rats showed a marked hypo-reactivity to phenylephrine (PE; 1nM to 10 microM) compared with aortas harvested from sham shocked rats. Hem shocked rats also had increased levels of TNF-alpha mRNA in the liver (15-20 min after the end of bleeding) and enhanced plasma levels of 2,5-dihydroxybenzoic acid (2,5-DHBA; 6 +/- 2.2 microm), 2,3-dihydroxybenzoic acid (2,3-DHBA; 13 +/- 2.1 microm), both studied to evaluate OH(*) production. Electrophoretic mobility shift assay showed that liver NF-kappaB binding activity increased in the nucleus 10 min after the end of hemorrhage and remained elevated until the death of animals. Western blot analysis suggested that the levels of inhibitory IkappaBalpha protein in the cytoplasm became decreased at 5 min after the end of bleeding. IRFI-042, a vitamin E analogue (20 mg/kg intraperitoneally 2 min after the end of bleeding), inhibited the loss of IkappaBalpha protein from the cytoplasm and blunted the increase in NF-kappaB binding activity. Furthermore IRFI-042 increased survival time (117.8 +/- 6.51 min; p <.01) and survival rate (vehicle = 0% and IRFI-042 = 80%, at 120 min after the end of bleeding), reverted the marked hypotension, decreased liver mRNA for TNF-alpha, reduced plasma TNF-alpha (21 +/- 4.3 pg/ml), and restored to control values the hypo-reactivity to PE. Our results suggest that acute blood loss (50% of the estimated total blood volume over a period of 20 min) causes early activation of NF-kappaB, likely through an increased production of reactive oxygen species. This experiment indicates that NF-kappaB-triggered inflammatory cascade becomes early activated during acute hemorrhage even in the absence of resuscitation procedures.

Resuscitating effect of melanocortin peptides after prolonged respiratory arrest.

1. The resuscitating activity of melanocortin peptides (MSH-ACTH peptides) was tested in an experimental model of prolonged respiratory arrest. 2. Anaesthetized, endotracheally intubated rats subjected to a 5 min period of ventilation interruption, invariably died from cardiac arrest within 6-9 min of resumption of ventilation. 3. When resumption of ventilation was associated with the simultaneous intravenous (i.v.) injection of a melanocortin peptide (alpha-MSH or ACTH-(1-24)) (160 microg kg(-1) there was an almost immediate (within 1 min), impressive increase in cardiac output, heart rate, mean arterial pressure (+ 560% of the before-treatment value) and pulse pressure (+356% of the before-treatment value), with full recovery of electroencephalogram after 30-45 min. Blood gases and pH were normalized within 15-60 min after treatment, and all treated animals eventually recovered completely and survived indefinitely (= more than 15 days). 4. The same response was observed in adrenalectomized animals, as well as in animals pretreated with a beta1-adrenoceptor blocking agent (atenolol, 3 mg kg(-1), i.v.), or with an alpha1-adrenoceptor blocking agent (prazosin, 0.1 mg kg(-1), i.v.), or with an adrenergic neurone blocking agent (guanethidine, 10 mg kg(-1), intraperitoneally). 5. An effect quite similar to that produced by melanocortins was obtained with ouabain (0.1 mg kg(-1), i.v.); the antioxidant drug, glutathione (75 mg kg(-1), i.v.) also produced 100% resuscitation, but the effect was slower in onset. On the other hand, adrenaline (0.005 mg kg(-1), i.v.) was able to resuscitate only 1 out of 8 rats and dobutamine (0.02 mg kg(-1), i.v.) resuscitated 4 out of 8 rats; moreover, the effect of both catecholamines was much slower in onset than that of melanocortins and the initial, impressive stimulation of cardiovascular function was absent. 6. These results show that melanocortin peptides have a resuscitating effect in a pre-terminal condition produced in rats by prolonged asphyxia. This effect seems primarily due to the restoration of cardiac function, not mediated by catecholamines. These data also suggest that these peptides may have potential therapeutic value in conditions of transient cardiac hypoxia and re-oxygenation such as occur in coronary artery disease.

Role of neuronal and vascular Ca(2+)-channels in the ACTH-induced reversal of haemorrhagic shock.

1. In a rat model of volume-controlled haemorrhagic shock causing the death of all control (saline-treated) animals within 30 min, the intravenous (i.v.) bolus injection of ACTH-(1-24) at a dose of 160 micrograms kg-1 produced an impressive and sustained restoration of arterial pressure, pulse pressure and respiratory function, with 100% survival at the end of the observation period (2 h). 2. Both intracerebroventricular (i.c.v., 0.015-0.06 microgram kg-1) and i.v. (5 micrograms kg-1) pretreatment with the N-calcium channel blocker, omega-conotoxin GVIA, and i.v. (but not i.c.v.) pretreatment with the L-calcium channel blocker, nicardipine (125-500 micrograms kg-1) dose-dependently prevented the ACTH-induced shock reversal. 3. These results further indicate that the effect of ACTH in haemorrhagic shock may involve a neuronal link and the eventual restoration of vascular tone mediated by N- and L-type calcium channels, respectively.

Tumour necrosis factor-alpha as a target of melanocortins in haemorrhagic shock, in the anaesthetized rat.

The cytokine tumour necrosis factor-alpha (TNF-alpha) is involved (mostly through the activation of inducible nitric oxide synthase) in the pathogenesis of circulatory shock. On the other hand, melanocortin peptides are potent and effective in reversing haemorrhagic shock, both in animals (rat, dog) and in humans. This prompted us to study the influence of the melanocortin peptide ACTH-(1-24) on the blood levels of TNF-alpha in haemorrhage-shocked rats and on the in vitro production of TNF-alpha by lipopolysaccharide (LPS)-activated macrophages. Plasma levels of TNF-alpha were undetectable before starting bleeding and greatly increased 20 min after bleeding termination in saline-treated rats. In rats treated with ACTH-(1-24) the almost complete restoration of cardiovascular function was associated with markedly reduced levels of TNF-alpha 20 min after bleeding termination. On the other hand, ACTH-(1-24) did not influence TNF-alpha plasma levels in sham-operated, unbled rats. In vitro, ACTH-(1-24) (25-100 nM) dose-dependently reduced the LPS-stimulated production of TNF-alpha by peritoneal macrophages harvested from untreated, unbled rats. These results indicate that inhibition of TNF-alpha overproduction may be an important component of the mechanism of action of melanocortins in reversing haemorrhagic shock.

Influence of ACTH-(1-24) on free radical levels in the blood of haemorrhage-shocked rats: direct ex vivo detection by electron spin resonance spectrometry.

1. The influence of ACTH-(1-24) on the blood levels of highly reactive free radicals in haemorrhagic shock was studied in rats. 2. Volume-controlled haemorrhagic shock was produced in adult rats under general anaesthesia (urethane, 1.25 g kg-1 intraperitoneally) by stepwise bleeding until mean arterial pressure stabilized at 20-23 mmHg. Rats were intravenously (i.v.) treated with either ACTH-(1-24) (160 micrograms kg-1 in a volume of 1 ml kg-1) or equivolume saline. Free radicals were measured in arterial blood by electron spin resonance spectrometry using an ex vivo method that avoids injection of the spin-trapping agent (alpha-phenyl-N-tert-butylnitrone). 3. Blood levels of free radicals were 6490 +/- 273 [arbitrary units (a.u.) ml-1 whole blood, before starting bleeding, and 30762 +/- 2650 after bleeding termination (means +/- s.e. mean of the values obtained in all experimental groups). All rats treated with saline died within 30 min, their blood levels of free radicals being 35450 +/- 5450 a.u. ml-1 blood, 15 min after treatment. Treatment with ACTH-(1-24) produced a rapid and sustained restoration of arterial pressure, pulse pressure, heart rate and respiratory function, with 100% survival at the end of the observation period (2 h); this was associated with an impressive reduction in the blood levels of free radicals, that were 12807 +/- 2995, 10462 +/- 2850, 12294 +/- 4120, and 10360 +/- 2080 a.u. ml-1 blood, 15, 30, 60 and 120 min after ACTH-(1-24) administration, respectively. 4. These results provide a direct demonstration that (i) in haemorrhagic shock there is a rapid and massive production of highly reactive free radicals, and that (ii) the sustained restoration of cardiovascular and respiratory functions induced by the i.v. injection of ACTH-(1-24) is associated with a substantial reduction of free radical blood levels. It is suggested that ACTH-(1-24) prevents the burst of free radical generation during blood mobilisation and subsequent tissue reperfusion, and this may be an important component of its mechanism of action in effectively preventing death for haemorrhagic shock.

Adrenocorticotropin reverses vascular dysfunction and protects against splanchnic artery occlusion shock.

1. Tumour necrosis factor (TNF-alpha) is involved in the pathogenesis of splanchnic artery occlusion (SAO) shock. On the other hand, inhibition of TNF-alpha is an important component of the mechanism of action of melanocortins in reversing haemorrhagic shock. We therefore investigated the effects of the melanocortin peptide ACTH-(1 - 24) (adrenocorticotropin fragment 1 - 24) on the vascular failure induced by SAO shock. 2. SAO-shocked rats had a decreased survival rate (0% at 4 h of reperfusion, while sham-shocked rats survived for more than 4 h), enhanced serum TNF-alpha concentrations (755+/-81 U ml-1), decreased mean arterial blood pressure, leukopenia, and increased ileal leukocyte accumulation, as revealed by means of myeloperoxidase activity (MPO=9.4+/-1 U g-1 tissue). Moreover, aortic rings from shocked rats showed a marked hyporeactivity to phenylephrine (PE, 1 nM - 10 microM) (Emax and ED50 in shocked rats=7.16 mN mg-1 tissue and 120 nM, respectively; Emax and ED50 in sham-shocked rats=16.31 mN mg-1 tissue and 100 nM, respectively), reduced responsiveness to acetylcholine (ACh, 10 nM-10 microM) (Emax and ED50 in shocked rats=30% relaxation and 520 nM, respectively; Emax and ED50 in sham-shocked rats=82% relaxation and 510 nM, respectively) and increased staining for intercellular adhesion molecule-1 (ICAM-1). 3. ACTH-(1 - 24) [160 microg kg-1 intravenously (i.v.), 5 min after SAO] increased survival rate [SAO+ACTH-(1 - 24)=80% at 4 h of reperfusion], reversed hypotension, reduced serum TNF-alpha (55+/-13 U ml-1), ameliorated leukopenia, reduced ileal MPO (1.2+/-0.2 U g-1 tissue), restored the reactivity to PE, improved the responsiveness to ACh and blunted the enhanced immunostaining for ICAM-1 in the aorta. 4. Adrenalectomy only in part - but not significantly - reduced the ACTH-induced shock reversal, the survival rate of SAO+ACTH-(1 - 24) adrenalectomized rats being 60% at 4 h of reperfusion; and methylprednisolone (80 mg-1 i.v., 5 min after SAO) had a non-significant effect (10% survival) at 4 h of reperfusion. 5. The present data show that melanocortins are effective also in SAO shock, their effect being, at least in part, mediated by reduced production of TNF-alpha. Furthermore, they demonstrate, for the first time, that this inhibition is responsible for the adrenocorticotropin-induced reversal of vascular failure and leukocyte accumulation.

Haematological changes induced by the intravenous injection of CCK-8 in rats subjected to haemorrhagic shock.

In rats bled to invariably fatal haemorrhagic shock (mean arterial pressure = 18-24 mmHg), the prompt and sustained improvement of cardiovascular function, obtained with the i.v. injection of cholecystokinin octapeptide (CCK-8, 20 g/kg) is associated with a massive increase in the volume of residual circulating blood (0.69 +/- 0.12 ml/100 g b.w. in saline-treated rats; 1.61 +/- 0.09 ml/100 g b.w. in CCK-8-treated rats). The number of red cells/mm3 and the % Hb content is the same in CCK-8-treated and in control rats. So, in a condition of severe haemorrhage, otherwise incompatible with survival, the i.v. injection of CCK-8 not only induces an impressive increase in arterial pressure and in circulating blood volume, but also greatly improves tissue oxygenation.

Influence of TRH on regional blood flow and metabolic acidosis in a model of volume-controlled hemorrhagic shock in rats.

In anesthetized rats, massive bleeding to a severe condition of hemorrhagic shock (invariably leading to death within 30 min) was obviously associated with a dramatic decrease in tissue blood flow and with profound modifications of several blood parameters leading to metabolic acidosis: decrease in arterial and venous pH, bicarbonate and BE, decrease in arterial pCO2 and in venous pO2 and SO2, increase in arterial pO2, venous pCO2 and venous lactate. The i.v. bolus injection of protirelin tartrate (TRH-T, 4 mg/kg), which produces a prompt and sustained reversal of the shock condition, caused a rapid increase in venous pO2, pCO2 and SO2; on the other hand, arterial and venous pH, bicarbonate and BE continued to decrease--and venous lactate to increase during the first few minutes after treatment. However venous pCO2 and lactate, as well as arterial and venous pH, returned to the pre-bleeding values within 60 min after treatment. The data are in keeping with the TRH-T-induced improvement of circulatory and respiratory functions, with mobilization of the residual blood from its capillary pooling and consequent immission of acid metabolites into the blood stream.

Tolerance develops to the behavioural effects of ACTH-(1-24) during continuous i.c.v. infusion in rats, and is associated with increased hypothalamic levels of beta-endorphin.

In rats, the continuous infusion of ACTH-(1-24) into a brain lateral ventricle (0.5 micrograms/h in the volume of 1.11 microliters, for 7 days) caused a significant inhibition of the subsequent behavioural response to the acute intracerebroventricular injection of the same peptide. Tolerance developed to all the most typical signs of the ACTH-induced behavioural syndrome (grooming, stretching, yawning, penile erection, inhibition of food intake), and was associated with a significant increase in the hypothalamic levels of beta-endorphin immunoreactivity.

Ischemia- and reperfusion-induced arrhythmias are prevented by putrescine.

The influence of putrescine on cardiac arrhythmias induced by either permanent ligature of the left anterior coronary artery or heart reperfusion following a 5-min coronary occlusion was studied in anesthetized rats. Reperfusion-induced arrhythmias were significantly prevented by the i.v. injection of 150-200 mg/kg of putrescine, the survival rate being 100% in treated animals and 40% in controls. At a dose level of 200-300 mg/kg i.v., putrescine also significantly reduced the duration of ventricular tachycardia induced by permanent coronary occlusion. These findings show that putrescine significantly reduces the consequences of cardiac ischemia and reperfusion, probably as a consequence of its multiple stabilizing effects at the membrane level.

Dopamine D1 receptors are involved in the ACTH-induced reversal of hemorrhagic shock.

In an experimental model of volume-controlled hemorrhagic shock causing the death of all rats within 30 min, the intravenous (i.v.) bolus injection of the adrenocorticotropic hormone fragment 1-24 (ACTH-(1-24)) (160 micrograms/kg) induced a prompt and sustained improvement of cardiovascular and respiratory function, with 100% survival 2 h after treatment. Pretreatment with either haloperidol, 300 micrograms/kg i.v. (antagonist at dopamine D1 and D2 receptors), or (R)-(+)-8-chloro-2,3,4,5-tetrahydro-3-methyl-5-phenyl-1H-3- benzazepin-7-ol hemimaleate (SCH 23390), 50 micrograms/kg intraperitoneally (selective antagonist at dopamine D1 receptors), significantly inhibited the effect of ACTH-(1-24). A complete inhibition was produced by intracerebroventricular pretreatment with SCH 23390 (0.1 micrograms/rat). On the other hand, both i.v. and i.c.v. pretreatment with l-sulpiride (selective antagonist at dopamine D2 receptors) (25 mg/kg and 80 micrograms/rat, respectively) had only minor effects. These data suggest that the mechanism of the ACTH-induced reversal of hemorrhagic shock involves the activation of dopamine D1 receptors in the brain.