Safety, tolerability, and pharmacodynamics of an anti-interleukin-1α/ß dual variable domain immunoglobulin in patients with osteoarthritis of the knee: a randomized phase 1 study.

OBJECTIVE: To investigate the safety, tolerability, pharmacokinetics, and pharmacodynamics of ABT-981, a human dual variable domain immunoglobulin simultaneously targeting interleukin (IL)-1α and IL-1ß, in patients with knee osteoarthritis (OA). METHOD: This was a randomized, double-blind, placebo-controlled, single-center study of multiple subcutaneous (SC) injections of ABT-981 in patients with mild-to-moderate OA of the knee (NCT01668511). Three cohorts received ABT-981 (0.3, 1, or 3 mg/kg) or placebo every other week for a total of four SC injections, and one cohort received ABT-981 (3 mg/kg) or placebo every 4 weeks for a total of three SC injections. Assessment of safety and tolerability were the primary objectives. A panel of serum and urine biomarkers of inflammation and joint degradation were evaluated. RESULTS: A total of 36 patients were randomized (ABT-981, n = 28; placebo, n = 8); 31 (86%) completed the study. Adverse event (AE) rates were comparable between ABT-981 and placebo (54% vs 63%). The most common AE reported with ABT-981 vs placebo was injection site erythema (14% vs 0%). ABT-981 significantly reduced absolute neutrophil count and serum concentrations of IL-1α/IL-1ß, high-sensitivity C-reactive protein, and matrix metalloproteinase (MMP)-derived type 1 collagen. Serum concentrations of MMP-derived type 3 collagen and MMP-degraded C-reactive protein demonstrated decreasing trends with ABT-981. Antidrug antibodies were found in 37% of patients but were not associated with the incidence or severity of AEs. CONCLUSION: ABT-981 was generally well tolerated in patients with knee OA and engaged relevant tissue targets, eliciting an anti-inflammatory response. Consequently, ABT-981 may provide clinical benefit to patients with inflammation-driven OA.

Enhanced COMP catabolism detected in serum of patients with arthritis and animal disease models through a novel capture ELISA.

OBJECTIVE: The study aimed determining whether assessment of cartilage oligomeric matrix protein (COMP) degradation products could serve as a serological disease course and therapeutic response predictor in arthritis. METHODS: We generated a panel of monoclonal antibodies against COMP fragments and developed a novel capture enzyme-linked immunosorbent assay (ELISA) for detecting COMP fragments in patients with osteoarthritis (OA) and rheumatoid arthritis (RA). This test was also used to monitor COMP fragments in surgically-induced OA, collagen-induced arthritis (CIA), and tumor necrosis factor (TNF) transgenic animal models. RESULTS: Compared with a commercial COMP ELISA kit that detected no significant difference in COMP levels between OA and control groups, a significant increase of the COMP fragments were noted in the serum of OA patients assayed by this newly established ELISA. In addition, serum COMP fragment levels were well correlated with severity in OA patients and the progression of surgically-induced OA in murine models. Furthermore, the serum levels of COMP fragments in RA patients, mice with CIA, and TNF transgenic mice were significantly higher when compared with their controls. Interestingly, treatment with TNFα inhibitors and methotrexate led to a significant decrease of serum COMP fragments in RA patients. Additionally, administration of Atsttrin [Tang, et al., Science 2011;332(6028):478] also resulted in a significant reduction in COMP fragments in arthritis mice models. CONCLUSION: A novel sandwich ELISA is capable of reproducibly measuring serum COMP fragments in both arthritic patients and rodent arthritis models. This test also provides a valuable means to utilize serum COMP fragments for monitoring the effects of interventions in arthritis.

Glatiramer acetate (GA), the immunomodulatory drug, inhibits inflammatory mediators and collagen degradation in osteoarthritis (OA) cartilage.

OBJECTIVE: Glatiramer acetate (GA), the generic name for Copaxone, an immunomodulatory agent, has been shown to induce interleukin-1 receptor antagonist (IL-1Ra) production in macrophages. We therefore tested the effects of GA on the catabolic activities of osteoarthritis (OA) chondrocytes. DESIGN: Primary human chondrocytes and OA cartilage explants were utilized in this study. IL-1Ra, pro-matrix metalloproteinase-13 (proMMP-13) and prostaglandin E(2) (PGE(2)) were estimated in the cell culture supernatants and in vitro MMP-13 activity was measured using fluorogenic substrate. TaqMan Real-Time quantitative polymerase chain reaction (RT-qPCR) was performed to estimate relative expression levels of genes. RESULTS: GA treatment significantly increased transcription and production of sIL-1Ra (P=0.001) in both culture models. Furthermore, addition of GA (100 µg) inhibited: (1) spontaneous collagen degradation as assayed by CTX II enzyme-linked immunosorbent assay (ELISA) [mean CTX II (ng/g cartilage)] in control was 7.79 [95% confidence interval (CI) 2.57-13.02]-3.415 (95% CI 0.81-6.02) (P=0.0286); (2) spontaneous proMMP-13 secretion [mean MMP-13 (ng/g cartilage)] in control was 16.98 (95% CI 7.739-26.23)-6.973 (95% CI 1.632-12.31) (P=0.0286); (3) production of IL-1ß-induced inflammatory mediators such as nitric oxide (NO) [mean NO (µM)] in IL-1 cultures was 11.47 (95% CI 7.10-15.83)-0.87 (95% CI 0.18-1.56) (P=0.0022); and (4) recombinant MMP-13 in vitro activity (15-25%; P=0.004). CONCLUSIONS: These data suggest that GA effects may be due to upregulation of IL-1Ra as well as direct inhibition of MMP-13 activity. Based on these studies, we propose that GA has potential for disease modifying properties in OA and should be evaluated in vivo in animal studies.

[New views of modern medicine regarding treatment with stem cells, its practical and ethical consequences]. / Nové pohledy moderní medicíny na lécbu pomocí kmenových bunek, její praktické a etické dusledky.

Embryonic stem (ES) cells recently became the Holy Grail for the treatment of multiple diseases including cancer. A recent discovery of Israeli scientists however shows that ES cells can also become one of the causes triggering cancer. They base this hypothesis on a case of an Israeli boy suffering from Ataxia teleangiectasia, who developed cancer 4 years after ES transplantation. DNA analysis of the tumors showed that the malignant cells originated from the transplanted ES cells of the donors and not from the recipient's own cells. Given the fact that the therapeutic application of ES cells is still in the beginning, it is therefore necessary to thoroughly verify and test all the risks of their potential therapeutic use. This article also discusses in detail ethical and other aspects of ES cells applications, in particular the differences in points of view between Christianity and Judaism. Israeli Health to address these issues draws on traditional Judaism. Judaism defines an individual as a man if he cans a separate existence, in comparison with the human embryo, which is still directly dependent on his or her mother. Therefore, in matters of saving lives, including treatment options for improving the living, treatment with ES cells is tolerated Judaism as a clear preference for live and self-sustaining individual, before a cluster of cells. On the other hand, it is clear that all these therapies must have their basic rules and ethical reasons. In Israel, research on ESC and allowed it to those obtained in the course of unsuccessful fertility treatment by artificial insemination.

The COX-2 inhibitor market withdrawals and prescribing patterns by rheumatologists in patients with gastrointestinal and cardiovascular risk.

OBJECTIVE: To examine effects of the COX-2 inhibitor market withdrawals on NSAID utilization among patients at increased risk of gastrointestinal (GI) and cardiovascular (CV) toxicities. METHODS: A prospective cohort study was conducted using patients enrolled in the Consortium of Rheumatology Researchers of North America (CORRONA) Registry. The study population included rheumatoid arthritis (RA) and psoriatic arthritis (PsA) patients prescribed NSAIDs by rheumatologists from 1/1/2003 to 12/31/2005. Three cohorts were defined based on calendar year. The primary outcome assessed whether or not an NSAID gastroprotective strategy was prescribed. Secondary outcomes included rates of COX-2 inhibitor utilization and gastroprotective co-therapy utilization, stratified by the presence of cardiac and GI risk factors. RESULTS: NSAID gastroprotection utilization decreased from 65.1% in 2003 to 47.7% (p<0.001) in 2005. COX-2 inhibitor use decreased from 55.1% to 29.2% (p<0.001), whereas nonselective NSAIDs (nsNSAIDs) use increased from 50.2% to 73.9% (p=<0.01). Among patients with two or more risk factors for NSAID related GI bleeding, gastroprotection decreased from 74.4% in 2003 to 60.9% (p<0.01). For patients with two or more CV risk factors from 2003 to 2005, COX-2 inhibitor utilization decreased significantly, whereas nsNSAID utilization increased significantly. CONCLUSIONS: The COX-2 inhibitor withdrawals resulted in a rapid decline in NSAID gastroprotection prescribed by participating U.S. rheumatologists despite the availability of other gastroprotective options. Channeling toward nsNSAID use was widespread, including among patients at increased CV risk. Longer term follow-up is required to determine the clinical significance of these changes in NSAID prescribing, particularly for NSAID-related GI and CV-related toxicities.

Inhibition of ADAMTS-7 and ADAMTS-12 degradation of cartilage oligomeric matrix protein by alpha-2-macroglobulin.

OBJECTIVE: As we previously reported, ADAMTS-7 and ADAMTS-12, two members of ADAMTS (a disintegrin and metalloprotease with thrombospondin motifs) family, degrade cartilage oligomeric matrix protein (COMP) in vitro and are significantly induced in the cartilage and synovium of arthritic patients [Liu CJ, Kong W, Ilalov K, Yu S, Xu K, Prazak L, et al. ADAMTS-7: a metalloproteinase that directly binds to and degrades cartilage oligomeric matrix protein. FASEB J 2006;20(7):988-90; Liu CJ, Kong W, Xu K, Luan Y, Ilalov K, Sehgal B, et al. ADAMTS-12 associates with and degrades cartilage oligomeric matrix protein. J Biol Chem 2006;281(23):15800-8]. The purpose of this study was to determine (1) whether cleavage activity of ADAMTS-7 and ADAMTS-12 of COMP are associated with COMP degradation in osteoarthritis (OA); (2) whether alpha-2-macroglobulin (a(2)M) is a novel substrate for ADAMTS-7 and ADAMTS-12; and (3) whether a(2)M inhibits ADAMTS-7 or ADAMTS-12 cleavage of COMP. METHODS: An in vitro digestion assay was used to examine the degradation of COMP by ADAMTS-7 and ADAMTS-12 in the cartilage of OA patients; in cartilage explants incubated with tumor necrosis factor-alpha (TNF-alpha) or interleukin-1-beta (IL-1beta) with or without blocking antibodies; and in human chondrocytes treated with specific small interfering RNA (siRNA) to knockdown ADAMTS-7 or/and ADAMTS-12. Digestion of a(2)M by ADAMTS-7 and ADAMTS-12 in vitro and the inhibition of ADAMTS-7 or ADAMTS-12-mediated digestion of COMP by a(2)M were also analyzed. RESULTS: The molecular mass of the COMP fragments produced by either ADAMTS-7 or ADAMTS-12 were similar to those observed in OA patients. Specific blocking antibodies against ADAMTS-7 and ADAMTS-12 dramatically inhibited TNF-alpha- or IL-1beta-induced COMP degradation in the cultured cartilage explants. The suppression of ADAMTS-7 or ADAMTS-12 expression by siRNA silencing in the human chondrocytes also prevented TNF-alpha- or IL-1beta-induced COMP degradation. Both ADAMTS-7 and ADAMTS-12 were able to cleave a(2)M, giving rise to 180- and 105-kDa cleavage products, respectively. Furthermore, a(2)M inhibited both ADAMTS-7- and ADAMTS-12-mediated COMP degradation in a concentration (or dose)-dependent manner. CONCLUSION: Our observations demonstrate the importance of COMP degradation by ADAMTS-7 and ADAMTS-12 in vivo. Furthermore, a(2)M is a novel substrate for ADAMTS-7 and ADAMTS-12. More significantly, a(2)M represents the first endogenous inhibitor of ADAMTS-7 and ADAMTS-12.

Overview of benefit/risk of biological agents.

Targeted tumor necrosis factor-alpha antagonists, first approved by the FDA in 1998, have had a significant impact on the treatment of patients with rheumatoid arthritis. In general, the benefit/ risk ratio for these agents and the IL-1 receptor antagonist, anakinra, has been quite favorable. However, infrequent adverse events can be serious and require continued pharmacovigilance. Infections, particularly tuberculosis and less commonly fungal infections, are among the most serious adverse events, especially given delays in diagnosis due to subtle or atypical presentations. Questions have also arisen regarding whether anti-TNF-alpha agents increase the risk of lymphoma, a complicated issue confounded by the multiple risk factors for lymphoma in patients with rheumatoid arthritis and low observed incidence rates of lymphoma, requiring prolonged monitoring. Additional rare reported complications include systemic lupus erythematosus-like syndromes, congestive heart failure and demyelinating syndromes (including cases resembling progressive multifocal leukoencephalopathy). Ongoing post-marketing surveillance of these and other serious adverse events is necessary to determine the true incidence rates, and whether a reassessment of the overall risk-benefit of tumor necrosis factor-alpha antagonists will be required.

Nitric oxide sustains nuclear factor kappaB activation in cytokine-stimulated chondrocytes.

OBJECTIVE: In the current studies we have examined the effects of nitric oxide, and its redox derivatives peroxynitrite and S-nitrosothiol, S-nitrosocysteine, on nuclear factor kappaB (NF-kappaB) activation in cytokine-stimulated bovine chondrocytes. METHODS: The kinetics of NF-kappaB activation (p65 nuclear translocation) were assessed by immunofluorescence and immunoblot assays. RESULTS: We observed that the two nitric oxide redox species, peroxynitrite and S-nitrosocysteine, exert opposing effects on NF-kappaB activation. However, in lipopolysaccharide (LPS)/cytokine-stimulated chondrocytes (LPS, IL-1beta and TNF-alpha (LIT)) in the presence or absence of the NOS inhibitor L-NG-monomethyl arginine citrate (L-NMMA), the results indicate that nitric oxide causes persistent activation of NF-kappaB, most likely via generation of the free radical derivative peroxynitrite. CONCLUSION: The studies indicate that while nitric oxide is not required for immediate NF-kappaB activation in cytokine-stimulated chondrocytes, its effect is to sustain nuclear translocation of p65 and thereby provide a persistent "on signal" to NF-kappaB dependent gene transcription. Persistent activation of NF-kappaB may represent a mechanism by which nitric oxide sustains catabolic processes and promotes cartilage degeneration in osteoarthritis.

Can the procedure for prolapsing hemorrhoids (PPH) be done twice? Results of a porcine model.

BACKGROUND: The procedure for prolapsing hemorrhoids (PPH) is a new surgical method for the treatment of symptomatic hemorrhoids. In cases of recurrent prolapse, the performance of a second PPH may result in a ring of mucosa and submucosa between the two circular staple lines. In this study, we used a porcine model to assess whether PPH can be safely performed twice. METHODS: Five adult pigs underwent two PPH procedures in one session, leaving a ring of approximately 1 cm of mucosa between the two staple lines. One month later, the pigs were examined under anesthesia. The anal canal was assessed using the following four methods: (a) clinical examination, (b) evaluation of mucosal blood perfusion at different levels of the anal canal via a laser Doppler flow detector, (c) measurement of concentrations of hydroxyproline and collagen to check for fibrosis, and (d) histopathological examination. RESULTS: At the completion of the study period, all five pigs showed no clinical evidence of anorectal dysfunction. On examination under anesthesia 1 month after surgery, there was no evidence of anal stenosis in any of the pigs. The mean mucosal blood flow between the two staple lines did not differ significantly from the flow measured proximally and distally (394 vs 363 and 339 flow units, respectively; p = NS). The collagen levels, based on hydroxyproline concentration, were 81 mcg/mg between the staple lines, compared to 82 and 79 proximally and distally, respectively ( p = NS). There was no significant difference in degree of fibrosis, as assessed histopathologically, between specimens taken from the ring between the staple lines and specimens taken from the area external to the staple lines. CONCLUSIONS: The results of this porcine model suggest that a second synchronous PPH is feasible. A controlled experience involving human subjects is required to determine the safety and usefulness of this technique in cases of metachronous application for recurrent or residual hemorrhoids.

Variability in serotonin and enterochromaffin cells in patients with colonic inertia and idiopathic diarrhoea as compared to normal controls.

AIM: To evaluate differences in distribution, density and staining intensity of enterochromaffin cells (EC) and serotonin cells (SC) in the colonic mucosa of patients with colonic inertia (CI), idiopathic diarrhoea (ID) and a control group. METHODS: Three groups were studied: 19 patients' colons after subtotal colectomy for CI, and 17 patients' biopsies for diarrhoea (>3 bowel movements/day) with histological findings of normal mucosa (excluding microscopic, eosinophillic and collagenous colitis). The third group included 15 patients who underwent colonoscopy and biopsy for indications other than constipation, inflammatory bowel disease, diarrhoea or neoplasm (control group). Specimen blocks were obtained in each case from the right and left colon. Immunohistochemical staining for EC and SC were done on 4 micro m sections from Hollandes fixed, paraffin embedded tissues with primary rabbit antibody against chromagranin A or serotonin, and biotynylated secondary antibody and enzyme labelled streptavidin. RESULTS: The number of EC in the mucosa of the left colon in patients with CI (16.8 +/- 10.2) and ID (19.9 +/- 9.7) were significantly higher than they were on the right side (CI: 9.4 +/- 6.0, ID: 12.1 +/- 5.3). However, there were no significant differences between the left and right sides in the control group (L: 10.3 +/- 5.3; R: 13.4 +/- 7.6). Although the quantity of EC in the left colon in both patients with CI (P < 0.05) and ID (P < 0.01) were significantly higher than in the controls, there was no significant difference between CI and ID. In both the right and left colon, the percentage of EC with low positive density was significantly higher (P < 0.01) while those cells with moderate or low staining intensity were significantly lower in patients with CI than in either patients with ID or control group. In patients with CI, the quantity of SC in the mucosa of the left colon (12.1 +/- 6.4) was higher than in the right (CI: 7.9 +/- 3.6; control 4.6 +/- 3.3; ID 4.6 +/- 2.9) (P = 0.0057). In contrast there was no significant difference in SC in either the ID or control groups. The quantity of SC in both sides of the colon was significantly higher both in patients with CI as compared to the control group (P < 0.01) and patients with CI vs. patients with ID (L = P < 0.01; R = P < 0.05). There was a significantly positive correlation between the numbers of EC and SC in patients with CI (L: r = 0.5425, P < 0.05; R: r = 0.745, P < 0.01). CONCLUSION: In patients with CI, EC increases possibly due to an increase in SC. Conversely, in patients with ID, the EC increase results from peptides other than SC. Our results suggest that different aetiological factors contribute to ID and CI.

Altered serotonin immunoreactivities in the left colon of patients with colonic inertia.

BACKGROUND: Serotonin is an important positive regulator of colonic motility and transit. Its quantity and distribution in the left colon could be abnormal in patients with colonic inertia (CI) and contribute to the disease. AIM: To evaluate serotonin positive cells and immuno-reactivities in the mucosa, submucosa and muscularis propria of the left colon from patients with CI was compared to a control group. PATIENTS AND METHODS: Nineteen patients who underwent subtotal colecotomy for CI were assessed. The control group consisted of 15 patients who underwent left hemicolectomy for colonic cancer; histologically normal tissue specimens were used. Immunohistochemical staining for serotonin was performed. In the mucosa, the average number of serotonin positive cells per microscopic field (200 x ) was determined. The positively stained area (square pixels) in the mucosa, submucosa and muscularis propria per microscopic field (200 x ) was calculated utilizing a computer image analysis program. RESULTS: In the mucosa, both the number of serotonin positive cells and positively stained area were significantly higher in the patient group than in controls (P < 0.05). The difference in serotonin positive area in the submucosa in the CI group compared to the control group was not statistically significant. There was a very significant correlation between the serotonin positive area in the submucosa and muscularis propria in controls (r=0.65, p < 0.01), but not in patients with CI. CONCLUSION: The increased serotonin level in patients with CI may contribute to the disease or be an adaptive response to some other pathology. The lack of a positive correlation in serotonin levels between the submucosa and muscularis propria in CI patients suggests that the coordinated distribution of serotonin may be disrupted in CI.

Hypercoagulability: clinical assessment and treatment.

This review emphasizes pathophysiology, clinical features, assessment, and therapy for hypercoagulability. Risk factors that further increase clotting include obesity, recent surgery, pregnancy, and cancer. Clinical examples of coagulation abnormalities may occur from single or multiple abnormalities and include both inherited and acquired defects. Laboratory testing undertaken at the time of acute thrombosis is often inaccurate or difficult to interpret. Individuals are best tested when they are not taking anticoagulants. Treatment of patients with either inherited or acquired abnormalities usually requires heparin compounds followed by warfarin, but the length of therapy has not yet been settled. Asymptomatic individuals with underlying hypercoagulability may not require treatment except in clot-promoting situations such as trauma, pregnancy, recent surgery, or use of venous access devices. The detection of one abnormality may no longer suffice because multiple defects can be found frequently. In patients with clotting and underlying risk factors, such as malignancy, pregnancy, estrogens, or surgery, an assessment for hypercoagulability should be considered.

N7: a novel multi-modality therapy of high risk neuroblastoma (NB) in children diagnosed over 1 year of age.

BACKGROUND: The N7 protocol for poor-risk neuroblastoma uses dose-intensive chemotherapy (as in N6 protocol [Kushner et al.: J Clin Oncol 12:2607-2613, 1994] but with lower dosing of vincristine) for induction, surgical resection and 2100 cGy hyperfractionated radiotherapy for local control, and for consolidation, targeted radioimmunotherapy with 131I-labeled anti-GD2 3F8 monoclonal antibody and immunotherapy with unlabeled/unmodified 3F8 (400 mg/m2). PROCEDURE: The chemotherapy consists of: cyclophosphamide 70 mg/kg/d x 2 and a 72-hr infusion of doxorubicin 75 mg/m2 plus vincristine 2 mg/m2, for courses 1, 2, 4, and 6; and cisplatin 50 mg/m2/d x 4 and etoposide 200 mg/m2/d x 3, for courses 3, 5, and 7. 131I-3F8 is dosed at 20 mCi/kg, which is myeloablative and therefore necessitates stem-cell support. RESULTS: Of the first 24 consecutive previously untreated patients more than 1 year old at diagnosis, 22 were stage 4 and two were unresectable stage 3 with MYCN amplification. Chemotherapy achieved CR/VGPR in 21 of 24 patients. Twenty patients to date have completed treatment with 131I-3F8, and 15 patients have completed all treatment. With a median follow-up of 19 months, 18 of 24 patients remain progression-free. CONCLUSIONS: Major toxicities were grade 4 myelosuppression and mucositis during chemotherapy, and self-limited pain and urticaria during antibody treatment. Late effects include hearing deficits and hypothyroidism.

Activation of stress-activated protein kinase in osteoarthritic cartilage: evidence for nitric oxide dependence.

OBJECTIVE: We have demonstrated in bovine chondrocytes that nitric oxide (NO) mediates IL1 dependent apoptosis under conditions of oxidant stress. This process is accompanied by activation of c-Jun NH2-terminal kinase (JNK; also called stress-activated protein kinase). In these studies we examined activation of JNK in explant cultures of human osteoarthritic cartilage obtained at joint replacement surgery and we characterized the role of peroxynitrite to act as an upstream trigger. DESIGN: A novel technique to isolate chondrocyte proteins (<10% of total cartilage protein) from cartilage specimens was developed. It was used to analyse JNK activation by a western blot technique. To examine the hypothesis that chondrocyte JNK activation is a result of increased peroxynitrite, in vitro experiments were performed in which cultured chondrocytes were incubated with this oxidant. RESULTS: Activated JNK was detected in the cytoplasm of osteoarthritis (OA) affected chondrocytes but not in that of controls. In vitro, chondrocytes produce NO and superoxide anion. IL-1 (48 h), which induces nitric oxide synthase, resulted in an activation of JNK; this effect was reversed by N-monomethylarginine (NMA). TNFalpha treated chondrocytes at 48 h produce superoxide anion (EPR method). Exposure of cells to peroxynitrite led to an accumulation of intracellular oxidants, in association with JNK activation and cell death by apoptosis. CONCLUSION: We suggest that JNK activation is among the IL-1 elicited responses that injure articular chondrocytes and this activation of JNK is dependent on intracellular oxidant formation (including NO peroxynitrite). In addition, the extraction technique here described is a novel method that permits the quantitation and study of proteins such as JNK involved in the signaling pathways of chondrocytes within osteoarthritic cartilage.

Benign metastasizing leiomyoma: clinical, imaging, and pathologic correlation.

OBJECTIVE: We describe the clinical presentation and the radiographic and CT findings of benign metastasizing leiomyoma. CONCLUSION: Benign metastasizing leiomyoma is an asymptomatic disease characterized by well-defined, numerous, pulmonary lesions without a preponderant distribution.

Effect of pneumoperitoneal pressure on tumor dissemination and tumor recurrence at port-site and midline incisions.

Over the past several years numerous cases of port site tumor recurrence after laparoscopic resection of a cancerous tissue have been reported. Possible mechanisms for tumor seeding include tumor removal, contaminated instruments, pneumoperitoneum, and aerosolization of tumor cells. This experiment examined the relationship among trocar contamination, aerosolization, and tumor recurrence with increasing pneumoperitoneal pressure using a hamster model. Increased pneumoperitoneal pressure significantly increased both instrument contamination and tumor recurrence at midline and port site incisions. Interestingly, increasing pneumoperitoneal pressure had no significant effect on the number of aerosolized tumor cells. The results reaffirm that the use of a reduced pneumoperitoneum or gasless laparoscopy may significantly lower port site tumor recurrence.

Treatment of the alcohol intoxications: ethylene glycol, methanol and isopropanol.

Intoxications with ethylene glycol, methanol, and isopropanol are among the most common ingestions, in the treatment of which a nephrologist plays an important role. These three substances have the ideal characteristics for intervention by hemodialysis, and the three parent compounds and their metabolites are readily dialyzable. Two of the three substances, ethylene glycol and methanol, are metabolized to more toxic substances, so that an early treatment strategy that removes the parent compound or blocks its metabolism can prevent the development of many of the adverse events that are often seen in these ingestions. Fomepizole, an inhibitor of alcohol dehydrogenase, slows the metabolism of these substances and is now approved by the US Food and Drug Administration for use in ethylene glycol intoxication. The present review addresses recent advances in the diagnosis and treatment of intoxication with ethylene glycol, methanol and isopropanol.

Chemical syntheses and biological activities of lactam analogues of alpha-conotoxin SI.

Bicyclization represents an effective method for the introduction of conformational constraints into small, biologically important peptides. Several strategies have been developed for the preparation of bicyclic lactam analogues of alpha-conotoxin SI, a 13-residue peptide neurotoxin found in cone snail venom. Four analogues of the natural regioisomer of alpha-conotoxin SI were designed and synthesized, each with one of the two paired cysteines of the parent peptide being replaced by a side-chain lactam bridged glutamic acid/lysine pair. Solid-phase lactamization was studied to determine rates of formation of the two possible loops and to document the extent of dimerization and higher oligomerization. Radioligand binding assays were carried out on all synthesized peptides, including the naturally occurring two-disulfide form, in order to determine their affinities for nicotinic acetylcholine receptors (nAChRs). Replacement of the Cys(2)-Cys(7) loop of alpha-conotoxin SI with a lactam bridge resulted in complete loss of activity, whereas replacement of the Cys(3)-Cys(13) disulfide loop resulted in a approximately 60-fold reduction in affinity for one orientation and a approximately 70-fold increase in affinity for the other. The two active lactam analogues retain the selectivity exhibited by the naturally occurring peptide for the alpha/delta subunit of nAChRs, as judged by competition experiments with the curariform antagonist metocurine.

Nitric oxide synthase/COX cross-talk: nitric oxide activates COX-1 but inhibits COX-2-derived prostaglandin production.

It is recognized that there is molecular cross-talk between the inflammatory mediators NO and PGs that may regulate tissue homeostasis and contribute to pathophysiological processes. However, the literature is divided with respect to whether NO activates or inhibits PG production. In this study, we sought to determine whether conflicting observations could be accounted for by divergent effects of NO on the two cyclooxygenase (COX) isoforms. Exposure of resting macrophages to NO (30 microM) enhanced PGE2 release by 4. 5-fold. This enhancement was inhibited by indomethacin but not by the COX-2 selective inhibitor NS398. To separate the activation of phospholipase A2 and COX, we performed experiments using fibroblasts derived from COX-1-deficient or COX-2-deficient mice. These cells exhibit increased basal PG production, which is due to a constitutively stimulated cytosolic phospholipase A2 and enhanced basal expression of the remaining COX isozyme. The exposure of COX- 2-deficient cells to exogenous NO (10 microM) resulted in a 2.4-fold increase of PGE2 release above controls. Further studies indicated that NO stimulated PGE2 release in COX-2-deficient cells, without altering COX-1 mRNA or protein expression. In contrast, NO inhibited COX-2-derived PGE2 production in both LPS-stimulated macrophages and COX-1 knockout cells. This inhibition was associated with both decreased expression and nitration of COX-2. Thus, these studies demonstrate divergent effects of NO on the COX isoforms. The regulation of PGE production by NO is therefore complex and will depend on the local environment in which these pleiotropic mediators are produced.

Differential anti-inflammatory effects of immunosuppressive drugs: cyclosporin, rapamycin and FK-506 on inducible nitric oxide synthase, nitric oxide, cyclooxygenase-2 and PGE2 production.

OBJECTIVE AND DESIGN: Cyclosporin, FK-506 and rapamycin have similar but distinct modes of interaction with cyclophilins, calcineurins and transcription factors. These immunosuppressive drugs have also been shown to inhibit cytotoxic and inflammatory responses in macrophage. Therefore, we evaluated the mechanism of action of these drugs on iNOS and COX-2 expression by macrophages, the products of which (NO and PGE2) have cytotoxic and proinflammatory activities. MATERIALS AND METHODS: The murine macrophage cell line RAW 264.7 was grown as monolayer cultures. The effects of pharmacologically relevant concentrations of cyclosporin, rapamycin and FK-506 were evaluated in the presence and absence of lipopolysaccharide (LPS) which is a known inducer of iNOS and COX-2. Subsequently the expression of iNOS and COX-2 were analyzed by Western and Northern analysis. The production of NO and PGE2 were assayed by Greiss and RIA respectively. RESULTS: Cyclosporin (1-5 microg/ml) and rapamycin (1.0-10 nM) but not FK-506 (5-10 nM) inhibited both iNOS and COX-2 expression at mRNA level which led to significant inhibition of NO and PGE2 production. CONCLUSION: These studies characterize differential mechanistic capacity of the immunophilin-binding immunosuppressive drugs (comparable to hydrocortisone) to inhibit both iNOS and COX-2 expression. Inhibition of iNOS and COX-2 mRNA accumulation by cyclosporin and rapamycin seem to be distinct. These studies also highlight potential anti-inflammatory properties of these drugs in addition to their known immunosuppressive activity.