Comparing the effect of aromatase inhibitor (letrozole) + cabergoline (Dostinex) and letrozole alone on uterine myoma regression,a randomized clinical trial.

OBJECTIVE: To evaluate the effect of letrozole in combination with cabergoline and letrozole alone on regression of symptomatic uterine myomas in women of reproductive age. DESIGN: Randomized controlled clinical trial. SETTING: University hospital. PATIENTS: Ninety-one women of reproductive age were enrolled in the study and 88 women were eligible. Eight participants were excluded from the study. INTERVENTIONS: Eighty women of reproductive age with symptomatic myomas >4cm were evaluated in two groups. Participants in Group 1 received 2.5mg letrozole once daily and cabergoline 0.5mg/week from the first day of the menstrual cycle for 12 weeks, and participants in Group 2 received letrozole alone. MAIN OUTCOME MEASURES: Changes in uterine size and volume; myoma size, volume and number; and side effects of treatment. RESULTS: Overall, 76 patients completed the study. Compared with baseline values, mean uterine volume was reduced significantly in both groups (p=0.01), and there was no significant difference between groups (p=0.99). The mean number of dominant myomas was reduced significantly in both groups (p=0.03), with no significant difference between groups (p=0.6). The mean volume of myomas was reduced significantly in both groups (p=0.01), with no significant difference between groups (p=0.45). Although a significant decrease in number and volume of myomas was documented in each group (p<0.05), the intergroup analyses did not reveal significant differences between the two groups in terms of the change in number (p=0.28) and volume (p=0.96) of myomas. Headache was significantly more common in the letrozole+cabergoline group (nine vs two cases, p=0.02), but the two groups were comparable for the remaining minor side effects. CONCLUSION: This study showed that 12 weeks of treatment with letrozole with and without cabergoline improved the size and volume of the uterus and myomas, led to symptom improvement, and could be used for short-term treatment prior to surgery or fertility programmes. CONDENSATION: Condensation letrozole in combination with cabergoline in the management of uterine fibroids.

Comparative study of vaginal danazol vs diphereline (a synthetic GnRH agonist) in the control of bleeding during hysteroscopic myomectomy in women with abnormal uterine bleeding: a randomized controlled clinical trial.

OBJECTIVE: To compare the usefulness of vaginal danazol and diphereline in the management of intra-operative bleeding during hysteroscopy. DESIGN: Randomized controlled clinical trial. SETTING: University hospital. PATIENTS: One hundred and ninety participants of reproductive age were enrolled for operative hysteroscopy. Thirty women were excluded from the study. INTERVENTIONS: One hundred and sixty participants with submucous myomas were allocated at random to receive either vaginal danazol (200mg BID, 30 days before surgery) or intramuscular diphereline (twice with a 28-day interval). MAIN OUTCOME MEASURES: Severity of intra-operative bleeding, clarity of the visual field, volume of media, operative time, success rate for completion of operation and postoperative complications. RESULTS: Overall, 145 patients completed the study. In the danazol group, 78.1% of patients experienced no intra-operative uterine bleeding, and 21.9% experienced mild bleeding. In the diphereline group, 19.4% of patients experienced no intra-operative uterine bleeding, but mild, moderate and severe bleeding was observed in 31.9%, 45.8% and 2.8% of patients, respectively. The difference between the groups was significant (p<0.001). A clear visual field was reported more frequently in the danazol group compared with the diphereline group (98.6% vs 29.2%, p<0.001). The mean operative time was 10.9 min and 10.6 min in the danazol and diphereline groups, respectively (p=0.79). The mean volume of infused media was 2.0L in both groups (p=0.99). The success rate was 100% for both groups with no intra-operative complications. CONCLUSION: Both vaginal danazol and diphereline were effective in controlling uterine bleeding during operative hysteroscopy. However, vaginal danazol provided a clearer visual field.

'Anteroposterior placenta': a novel, simple sonographic feature indicating benign vaginal bleeding in early pregnancy.

This study proposes a sonographic feature of the placenta in association with benign vaginal bleeding. A total of 286 normal singleton pregnancies were examined for 'anteroposterior, AP, placenta', which was reported when the placenta was attached to both anterior and posterior walls of the uterus in sagittal transabdominal ultrasound scans. Pregnancies were followed up by week 20. AP placenta, vaginal bleeding and spontaneous abortion were documented in 61 (21.3%), 44 (15.4%) and 2 (0.7%) pregnant women, respectively. AP placenta was significantly more common in the group with vaginal bleeding (54.5% vs 1.3%, p < 0.001, odds ratio = 6.65 with a 0.95 confidence interval of 3.34-13.24). Abortions occurred only in patients with vaginal bleeding and no AP placenta (10% vs 0%; p = 0.20). In a normal clinical pregnancy with no known risk of miscarriage, the presence of an AP placenta usually forecasts a benign vaginal bleeding/spotting in first 20 weeks of gestation.

Reaching the theoretical resonance quality factor limit in coaxial plasmonic nanoresonators fabricated by helium ion lithography.

Optical antenna structures have revolutionized the field of nano-optics by confining light to deep subwavelength dimensions for spectroscopy and sensing. In this work, we fabricated coaxial optical antennae with sub-10-nanometer critical dimensions using helium ion lithography (HIL). Wavelength dependent transmission measurements were used to determine the wavelength-dependent optical response. The quality factor of 11 achieved with our HIL fabricated structures matched the theoretically predicted quality factor for the idealized flawless gold resonators calculated by finite-difference time-domain (FDTD). For comparison, coaxial antennae with 30 nm critical dimensions were fabricated using both HIL and the more common Ga focus ion beam lithography (Ga-FIB). The quality factor of the Ga-FIB resonators was 60% of the ideal HIL results for the same design geometry due to limitations in the Ga-FIB fabrication process.

Mapping local charge recombination heterogeneity by multidimensional nanospectroscopic imaging.

As materials functionality becomes more dependent on local physical and electronic properties, the importance of optically probing matter with true nanoscale spatial resolution has increased. In this work, we mapped the influence of local trap states within individual nanowires on carrier recombination with deeply subwavelength resolution. This is achieved using multidimensional nanospectroscopic imaging based on a nano-optical device. Placed at the end of a scan probe, the device delivers optimal near-field properties, including highly efficient far-field to near-field coupling, ultralarge field enhancement, nearly background-free imaging, independence from sample requirements, and broadband operation. We performed ~40-nanometer-resolution hyperspectral imaging of indium phosphide nanowires via excitation and collection through the probes, revealing optoelectronic structure along individual nanowires that is not accessible with other methods.

Intrinsically aligned chemo-mechanical functionalization of twin cantilever structures.

Mechanical oscillators became a main focus of research in recent years for potential applications in biomolecule detectors. We recently demonstrated the feasibility of a scheme based on twin cantilevers with a sensitivity down to the single molecule. This approach is extremely promising under the condition that the two terminals of the device can be functionalized with high selectivity and nanometric accuracy by linker molecules. Here we demonstrate a chemo-mechanical method to achieve the intrinsically aligned functionalization of two silicon surfaces, which can be separated by a gap controllable with nanometric precision. The chemical binding of the target molecules in the selected position is obtained through a cycloaddition reaction which exploits the reactivity of the freshly cleaved surfaces that form when the cantilever gap is created. The general validity of this approach is shown by the use in different chemical environments of two compounds with different reactive functional groups.

Lipoxin A4 levels in asthma: relation with disease severity and aspirin sensitivity.

BACKGROUND: Lipoxin (LX) A4, an endogenous anti-inflammatory eicosanoid, has been found to be low in patients with severe asthma. However, few studies also suggested more diminished LX A4 levels in aspirin-exacerbated respiratory disease (AERD) when compared with aspirin-tolerant asthma (ATA). It is, therefore, currently not clear whether the asthma severity or the presence of AERD has a primary role in the disturbed LX metabolism. OBJECTIVE: To detect LX A4 and 15-epi-LX A4 levels in asthma patients with and without AERD of comparable severity. METHODS: The study groups consisted of 22 subjects with AERD, 22 subjects with ATA and 10 volunteers without asthma and aspirin sensitivity. Whole-blood samples were stimulated with calcium ionophore, A23187 (5 x 10(-5) m) and A23187 (5 x 10(-5) m)+aspirin (10(-4) m). LX A4 and 15-epi-LX A4 levels were analysed by the enzyme immune assay method. RESULTS: Severe asthma patients in both AERD [0.5 (0.8)] ng/mL and ATA [0.5 (0.45) ng/mL] groups showed diminished generation for LX A4 to stimulation with A23187 in comparison with other severity degrees in their groups (P=0.02 and 0.046, respectively). LX A4 generation in both severe groups was comparable with each other (P>0.05). Although severe cases with AERD showed a diminished capacity to generate 15-epi-LX A4, this did not reach statistical significance. CONCLUSION: This study indicated that diminished LX A4 generation was unique to severe asthma phenotype regardless of comorbid aspirin sensitivity. Clinical Implications Lower LX A4 levels in severe asthma would suggest a possibility for LX analogues as future treatment options in these patients.

Activation of cell cycle regulatory proteins in the apoptosis of terminally differentiated oligodendrocytes.

There is increasing evidence that proteins normally involved in the cell cycle play a role in the regulation of neuronal apoptotic death following various insults. However, it is not clear if the same mechanisms regulate cell death of oligodendrocytes as well. In this study, we investigated the mechanism of ceramide-induced apoptosis in primary rat oligodendrocytes. We show that ceramide treatment initiates a cascade of biochemical events involving cell cycle regulatory proteins. Although at the time of induction of cell death the oligodendrocytes are postmitotic, activation of c-myc and translocation of Cdc25A into the nucleus can be demonstrated. Of particular interest are the findings of the up-regulation of PCNA and down-regulation of p21WAF1/CIP1 protein, an inhibitor of cell-cycle progression. The current results show that activation of regulatory cell-cycle proteins at the oligodendrocytes G1-S checkpoint may constitute a crucial step of the death pathway of oligodendrocytes.

Rationally designed strings of promiscuous CD4(+) T cell epitopes provide help to Haemophilus influenzae type b oligosaccharide: a model for new conjugate vaccines.

The age-related and T cell-independent immunological properties of most capsular polysaccharides limit their use as vaccines, especially in children under 2 years of age. To overcome these limitations, polysaccharide antigens have been successfully conjugated to a variety of carrier proteins, such as diphtheria toxoid or tetanus toxoid (TT) and the diphtheria mutant (CRM197) to produce very successful glycoconjugate vaccines. The increasing demand for new conjugate vaccines requires the availability of additional carriers providing high and long-lasting T helper cell immunity. Here we describe the design and construction of three recombinant carrier proteins (N6, N10, N19) constituted by strings of 6, 10 or 19 human CD4(+) T cell epitopes from various pathogen-derived antigens, including TT and proteins from Plasmodium falciparum, influenza virus and hepatitis B virus. Each of these epitopes is defined as universal in that it binds to many human MHC class II molecules. When conjugated to Haemophilus influenzae type b (Hib) oligosaccharide, these carriers elicit a potent anti-Hib antibody response in mice. In the case of the N19-Hib conjugate, this response is at least as good as that observed with CRM197-Hib, a conjugate vaccine currently used for mass immunization. We also show that some of the universal epitopes constituting the recombinant carriers are specifically recognized by two human in vitro systems, suggesting that T cell memory is provided by the selected epitopes. The data indicate that rationally designed recombinant polyepitope proteins represent excellent candidates for the development and clinical testing of new conjugate vaccines.

Release of cysteinyl leukotrienes with aspirin stimulation and the effect of prostaglandin E(2) on this release from peripheral blood leucocytes in aspirin-induced asthmatic patients.

BACKGROUND: The decrease in prostaglandin E(2) (PGE(2)) release due to aspirin (ASA)-induced cyclooxygenase inhibition and the increment in cysteinyl leukotriene (Cys-LT) release secondary to the removal of the inhibitory effect of PGE(2) on Cys-LT release have been suggested in the pathogenesis of aspirin-induced asthma (AIA). OBJECTIVE: In this study, we aimed to investigate the in vitro release of Cys-LT and to determine the effect of PGE(2) on Cys-LT release from peripheral blood leucocytes of patients with AIA after stimulation by ASA. PATIENTS AND METHODS: Patients with AIA (n = 13), patients with ASA-tolerant asthma (ATA) (n = 12) and healthy volunteers as controls (n = 13) were included to the study. ASA and PGE2 at three different concentrations were applied to the peripheral blood leucocytes of the study group, and Cys-LT levels following stimulants were assessed by enzyme immunoassay method. RESULTS: There was no difference in baseline Cys-LT levels between groups (AIA 353.4 +/- 55.5 pg/mL, ATA 354.7 +/- 40.3 pg/mL, and control group 368.5 +/- 30.2 pg/mL; P > 0.05). Though not present in other groups, the Cys-LT level of 453.6 +/- 70.0 pg/mL following ASA stimulation was higher than baseline in patients with AIA (P = 0.04). When PGE(2) was added to the ASA-stimulated samples of patients with AIA, Cys-LT levels were measured as 298.7 +/- 78.6 pg/mL, 279.8 +/- 79.9 pg/mL, and 243.4 +/- 51.3 pg/mL at PGE(2) 10(-7) m, 10(-6) m and 10(-5) m concentrations, respectively. These levels were lower than the ASA-stimulated Cys-LT values (P = 0.03, P = 0.01 and P = 0.01, respectively). The inhibitory effect of different PGE(2) concentrations on Cys-LT release was also present in patients with ATA and in controls. CONCLUSION: The increase in Cys-LT levels following ASA stimulation seems to be unique to AIA, which was not present in patients with ATA and in healthy controls. The inhibitory effect of PGE(2) on stimulated Cys-LT levels is another important finding to elucidate the role of PGE(2) in the pathogenesis of AIA.

A method for separation of heparin species from biological samples by ethanol precipitation of compounds solubilized in guanidine hydrochloride.

In this paper we describe a procedure to determine glycosaminoglycan and oligosaccharide composition of biological samples such as cell cultures or tissue explants. We demonstrate that heparin species of different molecular mass can be easily fractionated by sequential ethanol precipitation in 4.0 M guanidine hydrochloride. We studied by gradient polyacrylamide gel electrophoresis fractionation of standard heparin and heparin-derived oligosaccharides by anion-exchange chromatography on DEAE-Sephacel resin eluted by increasing concentration of guanidine hydrochloride. The use of guanidine salts followed by sequential precipitation by increasing ethanol concentration allowed recovery of heparin and heparin-derived oligosaccharides.

Nuclear localization of cystatin B, the cathepsin inhibitor implicated in myoclonus epilepsy (EPM1).

Cystatin B is an anti-protease implicated in myoclonus epilepsy, a degenerative disease of the central nervous system. In vitro, cystatin B interacts with and inhibits proteases of the cathepsin family. Confocal microscopy analysis of the subcellular localization of cystatin B and cathepsin B shows that, in vivo, the two proteins are concentrated in different cell compartments. In fact, cystatin B is found mainly in the nucleus of proliferating cells and both in the nucleus and in the cytoplasm of differentiated cells, while cathepsin B, in either case, is essentially cytoplasmic. However, colocalization of cystatin and cathepsin B is observed in the isolated cell matrix and in the nuclear scaffold of differentiated neuroblastoma cells but not of proliferating cells. This suggests that at least a fraction of cystatin B is bound to the protease in differentiated cells. The electron microscopy analysis of the cell matrix confirms the observation made with confocal microscopy. The cellular activity of cathepsin B was analyzed with a fluorogenic cytochemical assay. A fluorescent signal is observed in the cytoplasm of proliferating cells but is undetectable in the cytoplasm of differentiated cells, suggesting that cathepsin B is active mainly during the cell cycle. This result is consistent with the separate compartimentalization of cystatin B and cathepsin B that we have observed in growing cells.

Effect of combination of misoprostol and indomethacin on eicosanoid production in carrageenan-induced air pouch inflammation in rats.

The effect of single or combined administration of indomethacin and misoprostol on the exudate leukocyte count and thromboxane B2, a stable metabolite of thromboxane A2, and on the leukotriene B4 level, as cyclooxygenase and lipoxygenase metabolites of arachidonic acid, was investigated in acute carrageenan-induced air pouch inflammation in rats. Administration of indomethacin (0.25 to 4 mg/kg) 1 h before carrageenan given by the orogastric route reduced the exudate leukocyte count and thromboxane B2 level whereas it increased the exudate leukotriene B4 level dose dependently. Administration of misoprostol, a synthetic prostaglandin E1 analogue, (12.5 to 100 microg/kg) twice daily for two days before carrageenan given by the orogastric route increased the exudate leukocyte count. Combined misoprostol and indomethacin did not change the effect of indomethacin alone on exudate leukocyte count. Misoprostol, when used alone, decreased exudate thromboxane B2 level significantly. However, misoprostol did not change the exudate leukotriene B4 level, while its combination with indomethacin prevented the indomethacin-induced increase in exudate leukotriene B4 level. In conclusion, although misoprostol can be combined with non-steroidal anti-inflammatory drugs in many chronic inflammatory situations, our results indicate that misoprostol may also be combined with indomethacin in acute inflammation without producing any change on the antiinflammatory efficacy of indomethacin in rats.

Constitutive expression of interleukin-1beta (IL-1beta) in rat oligodendrocytes.

The RT-PCR analysis of RNA from progenitor and differentiated primary rat oligodendrocytes, and from the oligodendrocyte CG-4 cell line, shows the presence of the IL-1beta mRNA, the type I IL-1beta receptor and the IL-1 receptor accessory protein in these cells. In situ hybridization of a rat IL-1beta probe to primary progenitor and differentiated rat oligodendrocytes results in a positive signal. The double hybridization of the IL-1beta probe, together with an oligodendrocyte-specific differentiation marker, to sections of postnatal rat brain at different stages of differentiation is also positive. The double immuno-labelling technique utilized indicates coincidence of the signals on the brain slices. The results show that IL-1beta mRNA is constitutively expressed in rat brain oligodendrocytes from 1 day after birth onward. In agreement with this observation, CG-4 cells, primary progenitor and differentiated rat oligodendrocytes are positively stained by antibodies against IL-1beta. Postnatal brain slices from 1 and 4 day old and adult rats, labelled with a double immunofluorescence technique, are also stained by antibodies against IL-1beta. This signal coincides with that of antibodies against oligodendrocyte-specific surface markers. We conclude that IL-1beta is constitutively expressed in rat brain progenitor and differentiated oligodendrocytes.

Defibrotide activity in experimental frostbite injury.

The pathogenesis of frostbite injury has not been completely elucidated although the available evidence suggests it is an inflammatory reaction following reperfusion injury. Defibrotide given i.p. at 40 mg/kg/ day for three days to rabbits, the ears of which were subjected to frostbite, decreased the presence of inflammatory cells (mast cells -76%; neutrophils -40.4%) and increased prostaglandin I2 (PGI2) (as 6-Keto-PGF1 alpha) in the involved skin. Thromboxane A2 (TxA2) (as TxB2) was unaffected. These data strengthen the view that an inflammatory process is the underlying cause of frostbite injury and that Defibrotide is active in pathological situations involving an inflammatory process like in frostbite.

Eicosanoids and inflammatory cells in frostbitten tissue: prostacyclin, thromboxane, polymorphonuclear leukocytes, and mast cells.

The pathophysiology of cold injury is still controversial. An inflammatory process has been implicated as the underlying mechanism and certain anti-inflammatory substances such as ibuprofen and acetylsalicylic acid have been used in the clinical treatment of frostbite injury. It has been postulated that the progressive ischemic necrosis is secondary to excessive thromboxane A2 production, which upsets the normal balance between prostacyclin (prostaglandin I2) and thromboxane A2. It was aimed to clarify the pathophysiology of cold injury in this study. Twenty-one New Zealand White rabbits, each weighing 1.2 to 2.9 kg, were divided into control (n = 10) and frostbitten (n = 11) groups the randomly. The rabbit ears in the frostbitten group were subjected to cold injury, and the levels of thromboxane A2 (as thromboxane B2) and of prostaglandin I2 (as 6-keto-prostaglandin F1alpha) and the number of inflammatory cells (polymorphonuclear leukocytes and mast cells) were measured in normal and frostbitten skin of rabbit ears. The levels of 6-keto prostaglandin F1alpha and thromboxane B2, the stable metabolites of prostaglandin I2 and thromboxane A2, respectively, were increased in a statistically significant way (p < 0.002) by frostbite injury; however, thromboxane B2 increased more than 6-keto prostaglandin F1alpha. Polymorphonuclear leukocytes and mast cells, absent in normal skin, were present in the frostbitten skin. There was a statistically significant (p < 0.01) correlation between the time a rabbit ear was maintained at below -10 degrees C and skin survival and between the weights of rabbits and skin survival (p < 0.024). All these findings suggest that inflammation is involved in frostbite injury; a decrease in prostaglandin I2/thromboxane A2 ratio could be one of the factors leading to necrosis; the bigger the animal, the better its ability to counter frostbite.

Effects of chronic treatment with indomethacin at clinically relevant doses on intestinal tissue 6-keto prostaglandin F1 alpha and leukotriene B4 level in relation to gastroenteropathy.

This study investigated the effects of indomethacin at clinically relevant doses and its chronic usage on intestinal pathology, survival time and intestinal tissue 6-keto prostaglandin F1 alpha and leukotriene B4 level in rats during various periods with different doses. Indomethacin was administered ranging from 0.625 to 5 mg/kg. When used in doses of 0.625 and 1.25 mg/kg, indomethacin caused no apparent intestinal lesions or death during a treatment period of 30 days. On the other hand, all rats died in 7 days when 5 mg/kg of indomethacin was given. Mortality rate reached 53.3% in seven days in the group where 3.75 mg/kg indomethacin was given. The minimal dose of indomethacin, which induced intestinal ulcer and death, was 2.5 mg/kg. The main pathological findings were intestinal ulcers, but no macroscopic and microscopic changes were observed in the stomach. Intestinal tissue 6-keto prostaglandin F1 alpha and leukotriene B4 levels were quantified by enzyme immunoassay after homogenisation and extraction of tissue. In dose-dependent studies, only the dose of indomethacin, 3.75 mg/kg, significantly inhibited intestinal tissue 6-keto prostaglandin F1 alpha levels during seven days application period (197.39 +/- 24.26 vs 383.66 +/- 46.68 ng/g tissue, treatment vs control). 2.5 mg/kg of indomethacin caused no intestinal ulceration on 4th day, however, it significantly inhibited intestinal tissue 6-keto prostaglandin F1 alpha levels on 4th day in time-dependent studies (190.3 +/- 26.62 vs 383.66 +/- 46.68 ng/g tissue, treatment vs control). Neither dose-dependent nor time-dependent indomethacin administration changed intestinal tissue leukotriene B4 level. The results of this study indicated that indomethacin produced enteropathy rather than gastropathy when used chronically in clinically relevant doses in rats. Inhibition of prostaglandin synthesis, which was estimated by quantification of intestinal tissue 6-keto prostaglandin F1 alpha level, seemed not to be a prerequisite for its enteropathic effect.

Induction of intracellular ceramide by interleukin-1 beta in oligodendrocytes.

The sphingomyelin pathway has been implicated in mediating the effect of several extracellular agents leading to important biochemical and cellular changes. The aim of this investigation is to study interleukin-1 beta (IL-1 beta) signaling in oligodendrocytes. For this purpose, the CG4 oligodendrocyte cells were differentiated and incubated with IL-1 beta. This treatment induced a time- and dose-dependent increase of the endocellular ceramide. To mimic the effect of the elevation of endogenous ceramide, the CG4 cells were treated with the ceramide analogue C2-ceramide. Cell survival, measured with the MTT assay, showed that, by increasing the concentration of ceramide, up to 40% of CG4 cells were dying within 6 h, similar data were obtained with the primary differentiated oligodendrocytes. Condensation of chromatin, nuclear fragmentation, and formation of apoptotic bodies indicated that apoptosis was the cause of death. Surprisingly, long-term exposure (72 h) to increasing concentrations of IL-1 beta, which increases intracellular ceramide, did not induce oligodendroglial cell death. These results show that an increase of intracellular ceramide is not sufficient to induce apoptosis in oligodendrocytes and that IL-1 beta signaling through the ceramide pathway in these cells can mediate functions other than programmed cell death.

Synthetic alleles at position 121 define a functional domain of human interleukin-1 beta.

The non-conservative substitution of the tyrosine residue at position 121 of human interleukin-1 beta (IL-1 beta) generates protein mutants showing strong reduction of the capacity to induce (a) prostaglandin E2 (PGE2) release from fibroblasts and smooth muscle cells, (b) murine T-cells proliferation and (c) activation of interleukin-6 (IL-6) gene expression. It is generally accepted that these functions are mediated by the type-I interleukin-1 receptor (IL-1RI). However, the mutant proteins maintain the binding affinity to the types-I and II IL-1 receptors, which is the same as the control IL-1 beta, suggesting that this amino acid substitution does not alter the structure of the molecule, except locally. Thus we have identified a new functional site of IL-1 beta different from the known receptor binding region, responsible for fundamental IL-1 beta functions. Moreover, we show that the same mutants maintain at least two hypothalamic functions, that is, the in vitro short-term PGE2 release from rat hypothalamus and the induction of fever in rabbits. This result suggests that there is yet another site of the molecule responsible for the hypothalamic functions, implying that multiple active sites on the IL-1 beta molecule, possibly binding to more than one receptor chain, trigger different signals.