Complementary and alternative medicine: the role of the cancer center.
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Adult↗
Biomedical subjects
J Chabot
Publications and source records attributed to J Chabot.
Interaction of gamma 1-syntrophin with diacylglycerol kinase-zeta. Regulation of nuclear localization by PDZ interactions.
Syntrophins are modular adapter proteins that link ion channels and signaling proteins to dystrophin and its homologues. A yeast two-hybrid screen of a human brain cDNA library using the PDZ domain of gamma 1- syntrophin, a recently identified brain-specific isoform, yielded overlapping clones encoding the C terminus of diacylglycerol kinase-zeta (DGK-zeta), an enzyme that converts diacylglycerol into phosphatidic acid. In biochemical assays, the C terminus of DGK-zeta, which contains a consensus PDZ-binding motif, was found to be necessary and sufficient for association with gamma 1-syntrophin. When coexpressed in HeLa cells, DGK-zeta and gamma 1-syntrophin formed a stable complex that partitioned between the cytoplasm and nucleus. DGK-zeta translocates from the cytosol to the nucleus, a process negatively regulated by protein kinase C phosphorylation. We found that DGK-zeta recruits gamma 1-syntrophin into the nucleus and that the PDZ-binding motif is required. Disrupting the interaction altered the intracellular localization of both proteins; DGK-zeta accumulated in the nucleus, whereas gamma 1-syntrophin remained in the cytoplasm. The level of endogenous syntrophins in the nucleus of HeLa cells also reflected the amount of nuclear DGK-zeta. In the brain, DGK-zeta and gamma 1-syntrophin were colocalized in cell bodies and dendrites of cerebellar Purkinjie neurons and other neuronal cell types, suggesting that their interaction is physiologically relevant. Moreover, coimmunoprecipitation and pull-down experiments from brain extracts and cells suggest that DGK-zeta, gamma 1-syntrophin, and dystrophin form a ternary complex. Collectively, our results suggest that gamma 1-syntrophin participates in regulating the subcellular localization of DGK-zeta to ensure correct termination of diacylglycerol signaling.
Adaptor Proteins, Signal Transducing↗
Single-amino acid substitutions alter the specificity and affinity of PDZ domains for their ligands.
PDZ domains are modular protein-protein interaction domains that bind to specific C-terminal sequences of membrane proteins and/or to other PDZ domains. Certain PDZ domains in PSD-95 and syntrophins interact with C-terminal peptide ligands and heterodimerize with the extended nNOS PDZ domain. The capacity to interact with nNOS correlates with the presence of a Lys residue in the carboxylate- binding loop of these PDZ domains. Here, we report that substitution of an Arg for Lys-165 in PSD-95 PDZ2 disrupted its interaction with nNOS, but not with the C terminus of the Shaker-type K(+) channel Kv1.4. The same mutation affected nNOS binding to alpha1- and beta1-syntrophin PDZ domains to a lesser extent, due in part to the stabilizing effect of tertiary interactions with the canonical nNOS PDZ domain. PDZ domains with an Arg in the carboxylate-binding loop do not bind nNOS; however, substitution with Lys or Ala was able to confer nNOS binding. Our results indicate that the carboxylate-binding loop Lys or Arg is a critical determinant of nNOS binding and that the identity of this residue can profoundly alter one mode of PDZ recognition without affecting another. We also analyzed the effects of mutating Asp-143, a residue in the alphaB helix of alpha1-syntrophin that forms a tertiary contact with the nNOS PDZ domain. This residue is important for both nNOS and C-terminal peptide binding and confers a preference for peptides with a positively charged residue at position -4. On this basis, we have identified the C terminus of the Kir2.1 channel as a possible binding partner for syntrophin PDZ domains. Together, our results demonstrate that single-amino acid substitutions alter the specificity and affinity of PDZ domains for their ligands.
Amino Acid Sequence↗
Endothelial-monocyte activating polypeptide II, a novel antitumor cytokine that suppresses primary and metastatic tumor growth and induces apoptosis in growing endothelial cells.
Neovascularization is essential for growth and spread of primary and metastatic tumors. We have identified a novel cytokine, endothelial-monocyte activating polypeptide (EMAP) II, that potently inhibits tumor growth, and appears to have antiangiogenic activity. Mice implanted with Matrigel showed an intense local angiogenic response, which EMAP II blocked by 76% (P < 0.001). Neovascularization of the mouse cornea was similarly prevented by EMAP II (P < 0.003). Intraperitoneally administered EMAP II suppressed the growth of primary Lewis lung carcinomas, with a reduction in tumor volume of 65% versus controls (P < 0.003). Tumors from human breast carcinoma-derived MDA-MB 468 cells were suppressed by >80% in EMAP II-treated animals (P < 0.005). In a lung metastasis model, EMAP II blocked outgrowth of Lewis lung carcinoma macrometastases; total surface metastases were diminished by 65%, and of the 35% metastases present, approximately 80% were inhibited with maximum diameter <2 mm (P < 0.002 vs. controls). In growing capillary endothelial cultures, EMAP II induced apoptosis in a time- and dose-dependent manner, whereas other cell types were unaffected. These data suggest that EMAP II is a tumor-suppressive mediator with antiangiogenic properties allowing it to target growing endothelium and limit establishment of neovasculature.
Animals↗
Evidence for tonic activation of NK-1 receptors during the second phase of the formalin test in the Rat.
Behavioral, electrophysiological, and autoradiographic experiments were done to study the second nociceptive phase in the formalin test. In initial experiments, this second phase was attenuated by 1-10 mg of the NK-1 receptor antagonist CP-99,994, given subcutaneously 10, 30, or 60 min before formalin (n = 8-10) and by 20 microgram given intrathecally 20 min after formalin (n = 13); the inactive isomer CP-100,263 was ineffective. In electrophysiological experiments on single dorsal horn neurons in vivo, the excitatory responses to subcutaneous formalin injection (50 microliter, 2.5%) were attenuated by subsequent intravenously administration of the NK-1 receptor antagonist CP-96,345 (0.5 mg/kg; n = 8), given 35-40 min after formalin, but not by the inactive enantiomer CP-96,344 (0.5 mg/kg; n = 9). Finally, autoradiographic binding of exogenous [(125)I]BH-substance P in the lumbar cord was reduced at 5 and 25 min after formalin (50 microliter, 1 or 5%), with an intermediate level of reduction at 12 min. These data are interpreted as evidence that the second phase of nociceptive scores in the formalin test is attributable at least partially to tonic activation of NK-1 receptors at the spinal level, whether because of a temporally limited release of substance P, for example only during the first phase, but a slow removal or breakdown of substance P, or, more likely, because of tonic release from primary afferents throughout the second phase. Irrespective of the mechanism, it can be concluded that at least some of the persistent nociceptive effects associated with peripheral inflammation, or at least those provoked by subcutaneous injection of formalin, are mediated via continuous activation of NK-1 receptors at the level of the spinal dorsal horn; this may relate directly to mechanisms underlying prolonged nociceptive pains in humans.
Animals↗
Long-term survival of a young woman with peripheral cholangiocarcinoma: a case report.
Cholangiocarcinoma is the second most common primary tumor of the liver after hepatocellular carcinoma and accounts for 5 to 25% of primary hepatic malignancies. Patients with intrahepatic or peripheral cholangiocarcinoma (ICC) most often present at an advanced stage leading to a poor prognosis. A review of the literature has produced only 10 patients who have survived over five years. We review the case of a young woman with a large cholangiocarcinoma, who has been disease free for eight years. The patient was treated with a right hepatic lobectomy, and received 4 cycles of 5-fluorouracil and levamisole postoperatively. Known factors associated with longer survival in patients with ICC include lack of evidence of local invasion (i.e. capsular, lymphatic, or vascular), negative margins, mucoblia, and well differentiation of the tumor, as well as the absence of lymph node metastases. Our patient had negative margins and lymph nodes, and showed no local invasion. However, no mucobilia was noted, and the tumor was only moderately differentiated. Young age has never been associated with increased survival. ICC remains a relatively uncommon tumor with an insidious onset and late presentation contributing to poor survival. Surgical resection remains the only therapeutic option. Since few patients are potentially resectable at the time of presentation, efforts at early diagnosis and options for adjuvant therapy are imperative.
Adult↗
Extracellular matrix induced by TGFbeta impairs insulin signal transduction in 3T3-L1 preadipose cells.
When 3T3-L1 preadipose cells are exposed to transforming growth factor beta (TGFbeta), they synthesize more extracellular matrix (ECM) and resist differentiation-inducing stimuli. The mechanism by which ECM suppresses adipose cell differentiation (adipogenesis) remains unknown. Since adipogenesis is an insulin/insulin-like growth factor-1 (IGF-1)-dependent process, we investigated whether TGFbeta-induced ECM inhibits insulin signaling. When preadipose cells were pretreated overnight with TGFbeta, we observed a 75% decrease in insulin-stimulated tyrosine phosphorylation of insulin receptor substrate-1 (IRS-1) compared to that in control cells. Culturing 3T3-L1 preadipose cells on fibronectin, a component of the ECM induced by TGFbeta, also inhibited insulin-dependent IRS-1 tyrosine phosphorylation and adipogenesis, supporting a role for ECM in mediating TGFbeta's inhibitory effect on insulin signaling. Since the insulin-stimulated association of phosphoinositide (PI) 3-kinase with IRS-1 depends on IRS-1 tyrosine phosphorylation, we measured the presence of the PI 3-kinase 85 kDa regulatory subunit in anti-IRS-1 immunoprecipitates. Following insulin stimulation, PI 3-kinase-IRS-1 association was reduced by 70% in TGFbeta pretreated vs. control preadipose cells. However, insulin-stimulated cellular production of PI(3,4,5)P3 was unaltered by TGFbeta pretreatment. This suggests that IRS-1-associated p85-type PI 3-kinase may represent a particular subset of total cellular PI 3-kinase that is specifically inhibited by TGFbeta. Reduction of insulin-stimulated association of IRS-1 with p85-type PI 3-kinase by TGFbeta may be one potential mechanism through which TGFbeta blocks 3T3-L1 adipose cell differentiation.
3T3 Cells↗
Liver metastases are enhanced in homozygous deletionally mutant ICAM-1 or LFA-1 mice.
BACKGROUND: Adhesion molecules play an integral role in tumor growth, invasion, and metastasis and have been shown to influence the immune response to malignant cells. The interaction of intercellular adhesion molecule-1 (ICAM-1) with lymphocyte function antigen-1 (LFA-1) is important for the adhesion of leukocytes, monocytes and lymphocytes to endothelial cells in vitro and in vivo. In order to explore the role of the ICAM-1/LFA-1 interaction in liver metastases, we utilized homozygous deletionally mutant (gene knockout) mice for ICAM-1 or LFA-1 which had been derived from the C57BL6/J background. MATERIALS AND METHODS: Wild-type C57BL6/J mice were used as controls. Animals were anesthetized and underwent a 1-cm midline lower abdominal incision. The ileocolic vein was identified and B16 melanoma cells (10(4)) were injected. The incisions were closed with skin clips. Two weeks following surgery, mice were sacrificed and their livers resected for gross and histological analysis. RESULTS: LFA-1 deficient mice developed 13 times the number of metastases compared to wild-type controls and ICAM-1 deficient mice developed 7 times that number [13.5 (n = 17) vs 1.0 (n = 19) and 36 (n = 10) vs 5.0 (n = 16), P values of 0.0003 and 0.0002 by Wilcoxon Rank Sum Test, respectively]: Histologically, multiple areas of inflammatory cells consisting of T-cells and macrophages were noted in wild-type mice. Only sparse inflammatory cells were noted surrounding the metastases in the null mice. CONCLUSIONS: Liver metastases of the B16 melanoma are markedly enhanced in ICAM-1 null and LFA-1 null mice. The ICAM-1/LFA-1 interaction is crucial to the immune response to liver metastases.
Animals↗
A metastasizing model of anaplastic human Wilms tumor in the nude mouse.
Reproducible animal models of Wilms tumor have been difficult to establish. We describe a model in which cells, banked from a patient with metastatic Wilms tumor, were implanted into nude mice, resulting in the development of primary renal and metastatic pulmonary lesions. Pathologically, the lesions resembled the blastemal component of anaplastic Wilms tumor. Primary tumors showed a significant propensity for growth in the kidney as opposed to other organs. Pulmonary metastases, histologically similar to the primary lesions, were regularly observed. This represents the first reproducible model of anaplastic, metastasizing human Wilms tumor. This system may prove effective for the study of factors influencing growth and angiogenesis in aggressive variants of Wilms tumor.
Anaplasia↗
A novel tumor-derived mediator that sensitizes cytokine-resistant tumors to tumor necrosis factor.
Therapeutic successes following treatment of murine tumors with tumor necrosis factor-alpha (TNF) have not been easily applied to clinical oncology because the concentrations of TNF required in humans induces systemic toxicity. This has led us to identify mediators which could sensitize tumors to the effects of TNF, permitting administration of lower doses and possible realization of the therapeutic potential of this cytokine. Our study reports the ability of a novel cytokine, endothelial-monocyte-activating polypeptide II (EMAP II), to sensitize initially resistant murine and human tumors to TNF-induced regression employing a murine model. Recombinant (r) EMAP II was purified from Escherichia coli transformed with a plasmid expressing mature EMAP II. The B16 melanoma, raised in C57BL/6 mice, or a human fibrosarcoma (HT-1080), grown in immunocompromised mice, was injected intratumorally with either vehicle or rEMAP II/heat-treated EMAP II (50-100 micrograms) followed by systemic TNF/heat-treated TNF (5 micrograms) and assessed for tumor volume, hemorrhage, and histologic appearance. Both the B16 melanoma and the HT-1080 human fibrosarcoma underwent thrombohemorrhagic and acute inflammatory changes concomitant with regression or significantly slowed growth after administration of intratumor EMAP II followed by systemic TNF. Omission or inactivation of either cytokine abrogated this effect. These results demonstrate that local treatment of certain tumors with EMAP II results in enhanced susceptibility to TNF-mediated induction of thrombohemorrhage and regression.
Animals↗
Parathyroid surgery using monitored anesthesia care as an alternative to general anesthesia.
BACKGROUND: Although there have been several reports in the literature describing a renewed interest in performing thyroid surgery under local anesthesia (LA), there has been little information regarding parathyroid surgery under local anesthesia. METHODS: We retrospectively reviewed our experience of 49 LA parathyroid patients over a 9-year period at a single institution. A bilateral cervical block (C2-C3) was administered by a single surgeon using lidocaine and bupivacaine. RESULTS: The study included 39 females and 10 males with an average age of 62 years (range, 35-89 years). Every surgery was curative and the final pathology revealed 46 parathyroid adenomas and 3 cases of parathyroid hyperplasia. Forty-seven percent of the patients were discharged within 6 hours of operation and the remaining patients had a 1.4-day average length of hospital stay. A group of age- and sex-matched controls who underwent parathyroid surgery using general anesthesia (GA) served as a control group with 27% of operations performed as outpatients and an average length of stay of 1.6 days. Return to work averaged 6 days for the LA group versus 8 days for the GA. In the LA group, there was one instance of postoperative hemorrhage requiring reoperation and one instance of conversion to GA secondary to an inability to tolerate LA. There were no instances of recurrent laryngeal nerve injury or permanent hypoparathyroidism in either group. CONCLUSIONS: These data suggest that experienced surgeons can perform parathyroid surgery safely and effectively using LA as an alternative to GA.
Adult↗
Detection of circulating thyroid cells in peripheral blood.
BACKGROUND: Detection of circulating malignant thyroid cells may provide a method to identify postoperative patients at risk for metastatic thyroid cancer. METHODS: On the basis of tissue specificity of thyroglobulin gene expression and the sensitivity of the reverse transcriptase-polymerase chain reaction (RT-PCR) analysis, we performed RT-PCR using primers for thyroglobulin on blood samples from patients with thyroid disease to detect thyroglobulin RNA transcripts. Postoperative peripheral blood samples from 100 patients, including patients with known metastatic thyroid cancer (six papillary and three follicular), thyroid cancer and no evidence of current metastases (63 papillary, 10 follicular, and five patients with both papillary and follicular), benign thyroid disease (six nontoxic nodular goiters), and normal volunteers (seven). RESULTS: Thyroglobulin transcripts were detected in nine of nine patients with metastatic thyroid cancer, seven of 78 patients with thyroid cancer and no current metastases (although of these seven patients, five had a history of metastatic disease that had been previously treated by surgery, one had a coexisting parathyroid cancer, and one had both papillary and follicular thyroid cancers), zero of six patients with benign thyroid disease, and zero of seven normal volunteers. Identity of amplicons was confirmed by restriction enzyme digestion and by cloning and sequencing of RT-PCR amplified thyroglobulin fragment (the latter in a limited number of cases). CONCLUSIONS: These data indicate that RT-PCR can be used to detect thyroglobulin mRNA in peripheral blood. The presence of these transcripts correlates with the existence of extrathyroidal disease.
Base Sequence↗
Expression of a constitutively activated form of protein kinase B (c-Akt) in 3T3-L1 preadipose cells causes spontaneous differentiation.
Insulin/IGF-1 is required for differentiation of 3T3-L1 adipose cells. Downstream targets of insulin/IGF-1 that lead to adipocyte differentiation appear to include Ras, phosphatidylinositol (PI) 3-kinase, Raf, and mitogen-activated protein kinase. We have tested whether protein kinase B (PKB), a serine/threonine kinase activated by PI 3-kinase, is sufficient for 3T3-L1 preadipose cell differentiation. A plasmid vector encoding a version of PKB that is constitutively activated (Gag-PKB) was expressed in 3T3-L1 preadipose cells (Gag-PKB cells). Spontaneous morphological changes indicative of adipocyte differentiation were observed in Gag-PKB cells. The cells assumed a spherical shape and they acquired characteristic lipid droplets that stained positively for Oil Red O. Northern blot analysis detected upregulation of LPL and aP2 mRNA, specific indicators of adipocyte differentiation. Our data demonstrate that constitutive activation of PKB is sufficient to trigger adipocyte differentiation.
3T3 Cells↗
Characterization of a novel tumor-derived cytokine. Endothelial-monocyte activating polypeptide II.
Endothelial-monocyte activating polypeptide II (EMAP II) was initially identified in the supernatant of murine methylcholanthrene A-induced fibrosarcomas (Meth A) by its capacity to activate host effector cells (Kao, J., Ryan, J., Brett, J., Chen, J., Shen, H., Fan, Y-G., Godman, G., Familletti, P., Wang, F., Pan, Y-C., Stern, D., and Clauss, M. (1992) J. Biol. Chem. 267, 20239-20247). Based on the NH2-terminal protein sequence, a full-length cDNA has been cloned which indicates that the precursor of EMAP II is a unique, leaderless, single polypeptide chain with predicted molecular mass approximately 34 kDa and that the mature form released by Meth A cells corresponds to approximately 20 kDa. Purified recombinant mature EMAP II (EMAP II, approximately 20 kDa form) activated endothelial cells with resulting elevation of cytosolic free calcium concentration, release of von Willebrand factor, induction of tissue factor, and expression of the adhesion molecules E-selectin and P-selectin. Neutrophils exposed to EMAP II demonstrated elevated cytosolic free calcium concentration, peroxidase generation, and chemotaxis. EMAP II also activated mononuclear phagocytes elevating cytosolic free calcium concentration, inducing tumor necrosis factor-alpha (TNF) and tissue factor, and stimulating chemotaxis. Systemic infusion of EMAP II into C3H/HeJ or Balb/c mice was associated with systemic toxicity, pulmonary congestion, and the appearance of TNF, interleukin-1 and -6 in the plasma. A single intra-tumor injection of EMAP II into Meth A sarcomas induced acute thrombohemorrhage and partial tumor regression. Local injection of EMAP II into a tumor resistant to the effects of TNF, murine mammary carcinoma, rendered it sensitive to subsequently administered TNF, which resulted in acute thrombohemorrhage and partial regression. These data suggest that recombinant EMAP II, a tumor-derived cytokine, has properties of a proinflammatory mediator with the capacity to prime the tumor vasculature for a locally destructive process.
Amino Acid Sequence↗
Thyroid surgery using monitored anesthesia care: an alternative to general anesthesia.
Forty consecutive patients undergoing thyroid surgery under local anesthesia (LA) by a single surgeon over a 5-year period were included in this retrospective review. In all cases, the indication for LA was patient request. The study included 29 females and 11 males with an average age of 44 years (range 22-66 years). Body habitus was thin in 12.5%, average in 67.5%, and obese in 20%. Operations consisted of 21 unilateral thyroid lobectomies, 3 partial thyroidectomies, 3 subtotal thyroidectomies, and 13 total thyroidectomies. The pathology revealed benign disease in 45% and malignant disease in 55%. All procedures were performed using lidocaine and/or bupivacaine to administer a deep cervical plexus block as well as a field block. Mild additional intraoperative intravenous sedation was provided in most cases. Two patients were converted emergently to general endotracheal anesthesia because of inability to tolerate LA in one and a seizure secondary to intraarterial injection of lidocaine in the other patient. There were no instances of wound infection hemorrhage, recurrent laryngeal nerve injury, or hypoparathyroidism. In conclusion, thyroid surgery in selected patients can be performed safely using LA by experienced surgeons. If patients are carefully prepared preoperatively, LA offers a simple and reasonable alternative to general anesthesia.
Adult↗
The role of passenger leukocytes in the immunogenicity of intestinal and cardiac allografts in the rat.
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Animals↗
The role of passenger leukocytes in the immunogenicity of intestinal and cardiac allografts in the rat.
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Animals↗
The intraoperative incidence of detectable bilateral and multicentric disease in papillary cancer of the thyroid.
This is a prospective study to determine the incidence of grossly detectable multicentric and bilateral cancer at operation in patients who, before surgery, were believed to have a unilateral lesion. The opposite lobe was inspected at the time of operation and the surgeon made a determination as to whether there was bilateral and/or multicentric disease present. A total thyroidectomy was then carried out and the accuracy of the surgeon's judgment was established by permanent-section pathologic examination. Fifteen of the 44 patients were assessed to have gross bilateral disease. Thirteen of these were confirmed by pathologic examination. One case of microscopic bilateral disease was not recognized by the surgeon. Seventeen patients were thought to have unilateral multicentric disease. Fifteen of these cases were confirmed by microscopy and an additional seven cases were documented to have secondary foci. It is obvious from this study that most disease that is called microscopic disease is actually palpable. A review of the literature confirms this. It was very unlikely for a patient who did not have unilateral multicentric disease to have a contralateral focus of carcinoma. The incidence of bilateral disease was 32%, and the incidence of multicentric disease was 50%; the surgeon was very accurate in assessing this. We believe that surgeons who advocate lobectomy as the primary treatment for thyroid cancer are recognizing grossly detectable disease in a significant number of patients and thus are doing total thyroidectomies in most patients with bilateral disease.
Adolescent↗