Activated STING in a vascular and pulmonary syndrome.

BACKGROUND: The study of autoinflammatory diseases has uncovered mechanisms underlying cytokine dysregulation and inflammation. METHODS: We analyzed the DNA of an index patient with early-onset systemic inflammation, cutaneous vasculopathy, and pulmonary inflammation. We sequenced a candidate gene, TMEM173, encoding the stimulator of interferon genes (STING), in this patient and in five unrelated children with similar clinical phenotypes. Four children were evaluated clinically and immunologically. With the STING ligand cyclic guanosine monophosphate-adenosine monophosphate (cGAMP), we stimulated peripheral-blood mononuclear cells and fibroblasts from patients and controls, as well as commercially obtained endothelial cells, and then assayed transcription of IFNB1, the gene encoding interferon-ß, in the stimulated cells. We analyzed IFNB1 reporter levels in HEK293T cells cotransfected with mutant or nonmutant STING constructs. Mutant STING leads to increased phosphorylation of signal transducer and activator of transcription 1 (STAT1), so we tested the effect of Janus kinase (JAK) inhibitors on STAT1 phosphorylation in lymphocytes from the affected children and controls. RESULTS: We identified three mutations in exon 5 of TMEM173 in the six patients. Elevated transcription of IFNB1 and other gene targets of STING in peripheral-blood mononuclear cells from the patients indicated constitutive activation of the pathway that cannot be further up-regulated with stimulation. On stimulation with cGAMP, fibroblasts from the patients showed increased transcription of IFNB1 but not of the genes encoding interleukin-1 (IL1), interleukin-6 (IL6), or tumor necrosis factor (TNF). HEK293T cells transfected with mutant constructs show elevated IFNB1 reporter levels. STING is expressed in endothelial cells, and exposure of these cells to cGAMP resulted in endothelial activation and apoptosis. Constitutive up-regulation of phosphorylated STAT1 in patients' lymphocytes was reduced by JAK inhibitors. CONCLUSIONS: STING-associated vasculopathy with onset in infancy (SAVI) is an autoinflammatory disease caused by gain-of-function mutations in TMEM173. (Funded by the Intramural Research Program of the National Institute of Arthritis and Musculoskeletal and Skin Diseases; ClinicalTrials.gov number, NCT00059748.).

Giant mucinous cystadenoma in a young patient. A case report.

The Authors describe the clinical case of a seventeen-year-old girl who presented with abdominalgia, fever, nausea and vomiting. During surgery it proved necessary to remove not only the appendix but also a voluminous mucinous cystadenoma of the ovary. The authors take this observation as a starting point for a description of the pathophysiological and clinical findings of these neoplasms.

[Hepatic hydatidosis: old and actual therapeutic strategies]. / L'idatidosi epatica: pregresse e attuali strategie terapeutiche.

The authors, reporting on their experience, confirm as the incidence of the hepatic hydatidosis and the complications related to this pathology today are very meaningful. They specify that the objectives to be pursued are: elimination of the parasite, prevention or treatment of the complications, prevention of the development of a new infestation. They dwell therefore upon the various techniques, both conservative and radical, that marked the history of the hepatic hydatidosis surgery, and they conclude affirming that the procedures of eradication of the parasite and the pericystectomy, in particular in laparotomic way, also represents the ideal treatment in consideration of the a little encouraging results of the medical therapy.

[Closed trauma of the spleen. Indications to surgical treatment]. / I traumi chiusi della milza. Indicazioni al trattamento chirurgico.

The Authors mention the historical evolution that led to consider the splenectomy as the ideal operation in patients with post-traumatic lesions of the spleen. They linger then on the actual knowledges about the physiopathology of this organ that determined a substantial change of mind to a conservative treatment, when possible. By reporting their experience of the last decade, they weigh up how every therapeutic choice must be consequent to an accurate clinical evaluation that suggests the surgical abdomen exploration in urgency or, on the contrary, the monitoring of the patient. They linger particularly on the validity of the investigation and quantization of organ lesions and of the subsequent hemoperitoneum. The Authors end affirming that an accurate diagnostic evaluation and the subsequent therapy must take place in a specialized Trauma Center that ensures intensive monitoring and, if necessary, a timely operation in these patients.

[Spontaneous splenic rupture in a patient with cytomegalovirus infection]. / Rottura spontanea di milza in presenza di infezione citomegalica. Descrizione di un caso clinico.

The Authors report a case of spontaneous splenic rupture in a patient with Cytomegalovirus infection, stress the main characteristics of this infection and describe the mechanism that, during the viral infection, causes morphological and functional alterations of the spleen. The hypersplenism secondary to hyperfunctioning, the formation of immunocomplexes with secondary infarction, mainly of the white pulp, and the disseminated intravascular coagulation are responsible, as in the observed case, of the rupture of splenic capsula. The Authors conclude that the wide diffusion of Cytomegalovirus infection needs the knowledge of all the pathologic signs of this infection to make a timely diagnosis and treatment.

Nosocomial infections in colo-rectal surgery of the old patient.

Every surgical act, especially in geriatric age, can be a relevant moment in the onset of nosocomial infections. This has a peculiar aspect in patient who undergo colo-rectal surgery, both in election and especially in emergency, in which the simple opening of intestines always involves a minimal contamination. In order to reduce the incidence of infections, and therefore the septic complications of this surgery, it is necessary to pay attention to the preparation of the surgical equipe, of the operating room, of the surgical instruments and, in election, to the careful preparation of the patient through a careful evaluation of the possible bio-umoral alterations, in order to correct them. The results of our experience allow us to say that the prevention of post-operatory sepsis find its main moment in the careful evaluation and eventual correction of the nutritional status, in the stimulation of the immune system, in the antibiotic prophylaxis both parenteral and topical, and, last but not least, in a correct surgical technique. All this is particularly important for patients affected by colo-rectal neoplastic and inflammatory diseases, for which the intestinal bacteria, more virulent in weak and fragile patients, often represent the source of contamination that can start a sepsis and then assume an important part in determining the final result of surgery.

Carboplatin-induced immune hemolytic anemia.

BACKGROUND: A case of acute hemolysis following therapy with carboplatin, an anticancer chemotherapeutic agent, was investigated. Hemolytic anemia has been associated with cisplatin, a related drug, but not with carboplatin. CASE REPORT: An 8-year-old boy was treated for an astrocytoma by monthly intravenous injections of carboplatin. Lower back pain was noted after 26 monthly injections, and overt intravascular hemolysis occurred after the 27th injection. The direct antiglobulin test was 4+ with anti-IgG and 1+ with anti-C3d. RESULTS: Blood samples obtained on Days 28 and 56 after the last injection were tested for carboplatin-dependent antibody. The direct antiglobulin test was 1+ with anti-IgG; the eluate was 1+ with and without carboplatin. The serum indirect antiglobulin test was negative in the absence of carboplatin, 3+ to 4+ in the presence of carboplatin, and 1+ with carboplatin-coated cells. Day 56 serum antibody titer was 64 (agglutination at 37 degrees C), 512 (indirect antiglobulin test) in the presence of carboplatin, and 8 (indirect antiglobulin test) with carboplatin-coated cells. CONCLUSION: The findings indicate a carboplatin-induced antibody reacting in vitro by a complex mechanism combining elements of "immune complex," drug adsorption, and autoantibody mechanisms. Drug-dependent hemolysis is a previously unreported but potentially serious complication of carboplatin therapy.

Phase I trial of etoposide, doxorubicin and cisplatin (EAP) in combination with GM-CSF.

The aim of this study was to ameliorate the toxicity of the etoposide, doxorubicin and cisplatin (EAP) regimen and to investigate the feasibility of dose escalation, using the molgramostim form of granulocyte macrophage-colony stimulating factor (GM-CSF) 10 micrograms/kg/day s.c. into the regimen. The design of the trial allowed for amended scheduling of the agents in the event of suboptimal results. Initially the regimen comprised etoposide 120 mg/m2, days 1-3, doxorubicin 40 mg/m2, day 1, and cisplatin 40 mg/m2, days 2 and 8. GM-CSF was begun on day 4 and continued until recovery of granulocyte counts. Courses were repeated every 21 days. 3 patients were treated at these doses. 5 patients received escalated doses (etoposide 180 mg/m2; doxorubicin 60 mg/m2; cisplatin 60 mg/m2) on this schedule; 4 out of 5 had intolerable myelosuppression (grade IV neutropenia or thrombocytopenia lasting > or = 7 days). These results prompted the administration of the day 8 cisplatin dose on day 3, with GM-CSF beginning on day 4. At the lowest doses of each agent (etoposide 120-doxorubicin 40-cisplatin 40), 3 of 6 patients had intolerable myelosuppression, and 3 patients had febrile neutropenia. Dose escalation of all of the drugs to etoposide 180 mg/m2, doxorubicin 60 mg/m2, cisplatin 60 mg/m2 resulted in documented infections in 4 out of 4 patients. GM-CSF toxicity included rash, dyspnoea, arrhythmias and pericardial effusions. The conclusion was that the use of GM-CSF does not permit escalation of drug doses on either schedule of EAP administration, and that these results do not support the combined use of GM-CSF and EAP.

Insulin-like growth factor II overexpression in myoblasts induces phenotypic changes typical of the malignant phenotype.

The objective of this study was to examine the role of insulin-like growth factor II (IGF-II) in the pathogenesis of human rhabdomyosarcomas (RMS). We have demonstrated previously that RMS express high levels of IGF-II mRNA, secrete IGF-II peptide, and express both IGF-I and IGF-II receptors. Moreover, we showed that IGF-II functions as an autocrine growth and motility factor in RMS. Since IGF-II is expressed at high levels in fetal muscle cells and RMS are tumors thought to derive from skeletal myoblasts arrested along the normal myogenic pathway, autocrine production of IGF-II by RMS may be an etiological event in the development of this tumor. We have developed a model system which enabled us to study the effects of endogenous IGF-II overprotection in muscle myoblasts. Human cDNA for pre-prohormone IGF-II was transfected into mouse myoblasts in order to achieve high, constant expression of this growth factor, which is normally down-regulated at the end of the differentiation process. Expression of high IGF-II levels resulted in: (a) an increased proliferative rate; (b) impairment of the ability to differentiate into myoblasts; and (c) acquisition of the capability of anchorage-independent growth. No changes in the expression of IGF-I receptors were noted. We conclude that IGF-II overexpression in muscle myoblasts induces morphological and biological changes typical of the malignant phenotype and represents a fundamental event in the pathogenesis of RMS and possibly of other embryonal tumors.

Signals transduced via insulin-like growth factor I receptor (IGF(R)) mediate resistance to retinoic acid-induced cell growth arrest in a human neuroblastoma cell line.

Retinoic Acid (RA) has been shown to control growth and induce differentiation in a number of human neuroblastoma (NB) cell lines. However, a number of NB cell lines may be termed resistant to RA as they fail to growth arrest and differentiate. In studying the mechanism mediating RA-resistance, we noted that invariably RA-resistant NB cell lines constitutively express Insulin-like Growth Factor 2 (IGF2) (Gaetano, 1991b). The NB cell line LAN-1-15N (15N) represented an interesting model in which to study the development of RA-resistance as initially 15N cells are growth arrested by RA, however with prolonged culture (8-10 days) cells begin to proliferate. Coincidentally, RA induces IGF2 mRNA and protein secretion in 15N NB cells (Matsumoto, 1992). In this study we isolated RA-resistant 15N cell lines and analyzed their growth properties and changes in cell cycle related (cdc2, cdk2, cyclins A, B, D and E) and early response (fos and jun) gene expression to evaluate the role IGF2 may play in mediating RA resistance. We found that exogenous IGF2 stimulates growth in 15N and is capable of altering RA induced inhibition of NB cell growth. Finally we show that by blocking the Insulin-like Growth Factor Receptor (IGF1(R)) with a monoclonal antibody (alpha-IR3) in the presence of RA the growth of RAR cell lines could be completely blocked. These data are consistent with the concept that signals by IGF2 and transduced via the IGF1(R) can mediate resistance to the growth inhibiting properties of RA.

Specific expression of insulin-like growth factor-II in rhabdomyosarcoma tumor cells.

Insulin-like growth factor II (IGF-II) acts as autocrine growth and motility factor in human rhabdomyosarcoma cell lines, and Northern blot analysis of tumor biopsy specimens from both alveolar and embryonal rhabdomyosarcoma demonstrates high levels of IGF-II mRNA expression. To determine the frequency and site of expression of IGF-II in these tumors, the authors performed in situ hybridization. All tumor specimens examined expressed the gene for IGF-II, and this expression was localized to the tumor cells themselves and not to the surrounding stroma. These data suggest that the IGF-II autocrine loop may be operating not only in vitro but also in vivo.

The insulin-like growth factor II (IGF-II)/mannose 6-phosphate receptor mediates IGF-II-induced motility in human rhabdomyosarcoma cells.

Insulin-like growth factor-II (IGF-II) is an autocrine growth and motility factor for human rhabdomyosarcoma. It interacts with three different receptors: the IGF-I, the IGF-II, and the insulin receptor. A specific function of the IGF-II receptor in mediating IGF-II responses has not been defined. In this report we investigate the mechanism of IGF-II-mediated motility in rhabdomyosarcoma cells. We demonstrate that IGF-II and [Leu27]IGF-II, an analog selective for the IGF-II receptor, stimulate motility at concentrations in which they interact only with their own receptor. An antibody that blocks the IGF-I receptor does not inhibit either peptide activity, while an antibody specific for the IGF-II receptor suppresses the IGF-II-induced motility. This antibody does not interfere with rhabdomyosarcoma cell proliferation. We conclude that in rhabdomyosarcoma cells IGF-II stimulates two different responses mediated by distinct receptors: 1) a mitogenic response through the type I receptor and 2) a motility response through the type II receptor.

Clopidogrel inhibits the binding of ADP analogues to the receptor mediating inhibition of platelet adenylate cyclase.

Clopidogrel, like the homologous thienopyridine derivative ticlopidine, selectively inhibits platelet aggregation induced by ADP. We have previously described two nucleotide-binding sites on platelets related to ADP-mediated platelet responses. The first is a high-affinity binding site for 2-methylthio-ADP (2-MeSADP) that is linked to the inhibition of stimulated adenylate cyclase. The second is the 100-kd exofacial membrane protein aggregin, which is labeled by the reactive ADP analogue 5'-p-fluorosulfonylbenzoyl adenosine (FSBA) that is related to shape change and aggregation. We set out to determine if either of these sites is blocked in vivo by clopidogrel or its active metabolite. Six subjects were given clopidogrel (75 mg/day for 10 days) in a double-blind crossover experiment. All of the subjects developed prolonged bleeding times while taking the drug. The rate of onset of the effect on bleeding time varied among subjects. Platelet aggregation induced by ADP or thrombin was significantly impaired by the drug treatment, but no effect was detected on shape change. The incorporation of [3H]FSBA into aggregin was also unaffected. Inhibition of adenylate cyclase by ADP or by 2-MeSADP was greatly reduced in all subjects, and in the case of 2-MeSADP, there was evidence for a noncompetitive effect. Inhibition of adenylate cyclase by epinephrine was unaffected. In the three subjects for whom binding measurements were made, the number of binding sites for [32P]2-MeSADP was reduced from 534 +/- 44 molecules per platelet during control and placebo periods (11 determinations) to 199 +/- 78 molecules per platelet during drug treatment (three determinations). There was no consistent change in the binding affinity.(ABSTRACT TRUNCATED AT 250 WORDS)

Suramin inhibits the growth of human rhabdomyosarcoma by interrupting the insulin-like growth factor II autocrine growth loop.

Suramin is a polysulfonated naphthyl-urea with antineoplastic activity that binds various peptide growth factors. Since we previously demonstrated that insulin-like growth factor II (IGF-II) is an autocrine growth factor in human rhabdomyosarcoma (RMS), we studied the effect of suramin on the growth of human RMS cells. Suramin caused a dose-dependent decrease of RMS cell number grown either in 10% fetal bovine serum or in serum-free medium (half-maximal effective dose in mitogenic assays, 1.6 x 10(-4) and 9 x 10(-5) M, respectively). IGF-II and IGF-I added to RMS cells in the presence of suramin reversed the suramin-induced inhibition of cell growth. Since IGF-II exerts its mitogenic effects on RMS cells by binding to the type I receptor, we performed radioreceptor assays using 125I-IGF-I and found that suramin displaced 125I-IGF-I from the type I IGF receptor. There was an excellent correlation between the doses of suramin effective in inhibiting the growth of RMS cells and those that displaced the binding of IGF-I. Our data indicate that suramin exerts its effect on RMS cell growth by interfering with the binding of IGF-II to the type I IGF receptor, thereby interrupting the IGF-II autocrine growth in these cells. Disrupting autonomous growth of RMS may be a promising novel therapeutic approach.

Insulin-like growth factor II acts as an autocrine growth and motility factor in human rhabdomyosarcoma tumors.

Rhabdomyosarcoma is the most common soft tissue sarcoma of childhood and appears to arise from developing striated muscle-forming cells. Since insulin-like growth factor II (IGF-II) is involved in normal muscle growth and maturation and elevated IGF-II mRNA levels have previously been reported in rhabdomyosarcomas, we have been studying the possible role of IGF-II in the unregulated growth and invasive potential of these embryonal tumors. In this study, we demonstrate that 13 of 14 rhabdomyosarcoma tumors express high levels of IGF-II mRNA relative to normal adult muscle and also express mRNA for the type I IGF receptors on their cell surface, the receptor thought to mediate the effects of IGF-II on muscle cells. We have established several rhabdomyosarcoma cell lines in mitogen-free media and demonstrate that these cells express type I IGF receptors on their cell surface and secrete IGF-II into the media. Exogenous IGF-II is able to stimulate cellular motility in these cell lines as assayed in a modified Boyden chamber. Finally, alpha IR-3, a type I receptor antagonist, inhibits the growth of these cell lines in serum-free media but does not inhibit IGF-II-induced motility of these cells. These data suggest that endogenously produced IGF-II functions as an autocrine growth and motility factor in many rhabdomyosarcoma tumors. The mitogenic actions of IGF-II are mediated through a domain of the type I IGF receptor that is blocked by alpha IR-3. IGF-II-induced motility may be mediated through an alternative signaling pathway.

Elimination of clonogenic malignant human T cells using monoclonal antibodies in combination with 2'-deoxycoformycin.

2'Deoxycoformycin (dCF) specifically inhibits adenosine deaminase (ADA) and causes selective cytotoxicity of normal and malignant T cells. In clinical trials, dCF caused rapid lysis of malignant T lymphoblasts. Although dCF has been associated with dose-limiting nonhematopoietic toxicities, myelosuppression has not been observed. Since dCF is relatively nontoxic to hematopoietic stem cells, we tested dCF for utility in the ex vivo purging of malignant T lymphoblasts from remission leukemic bone marrow for autologous bone marrow transplantation. We found that T lymphoblast cell lines were sensitive to dCF (plus deoxyadenosine [dAdo]) under conditions that did not ablate human hematopoietic colony-forming cells. Moreover, combined pharmacologic (dCF plus dAdo) and immunologic (anti-T cell monoclonal antibodies [McAb] plus complement) purging resulted in additive reduction in clonogenic T lymphoblasts. These results provide the basis for a clinical trial of bone marrow transplantation using combined pharmacologic/immunologic purging of T lymphoblasts from patients' harvested autologous marrow.

Hysteroscopic retrieval of occult IUD.

PIP: The direct view of the uterine cavity offered by hysteroscopy can provide valuable information about the position of an IUD, its condition, and its relationship with other formations, thereby facilitating its removal. The diagnostic and therapeutic reliability of this technique was analyzed in 8 women with occult IUDs. Involved were 3 Nova T IUDs, 2 Gravigards, 2 Minigravigards, and 1 Progestasert device. 1 patient had undergone unsuccessful curettage for removal of the IUD; none of the other women had experienced any surgical manipulation prior to the hysteroscopy. The surgery consisted of 4 steps: 1) distention of the uterine cavity by insufflation of carbon dioxide, 2) hysteroscopic location of the IUD, 3) sighted grasping of the IUD or IUD segment with forceps, and 4) removal of the device together with the hysteroscope under dynamic vision. Hysteroscopic location and removal were successful in all 8 patients, with no complications. Hysteroscopy is thus recommended as an alternative retrieval technique, especially in cases when it is necessary to assess the deterioration and possible myometrial penetration of the IUD or the inflammation of the uterine cavity.^ieng

T-lymphocyte subpopulation changes during hemolysis in glucose-6-phosphate dehydrogenase (G6PD)-deficient children.

Lymphocyte subpopulations were studied by means of sheep red blood cell (SRC)-rosetting technique and OKT11, OKT3, OKT4, and OKT8 monoclonal antibodies in ten Sicilian glucose-6-phosphate dehydrogenase-deficient children during the hemolytic crisis due to fava bean ingestion. The number of SRC-rosetting lymphocytes was significantly reduced, while the number of OKT3-positive cells was normal. An inversion of the OKT4/OKT8 ratio was observed, due to a decrease of OKT4-positive cells and an increase of OKT8-positive cells. All these abnormalities reverted to normal in the four children studied after recovery.