Candesartan cilexetil ameliorates NOSTRIN-NO dependent portal hypertension in cirrhosis and ACLF.

In decompensated cirrhosis, the severity of portal hypertension (PHT) is associated with increased hepatic endothelial nitric oxide synthase (eNOS) trafficking inducer (Nostrin), but the mechanism remains unclear. AIM: To investigate: (1) Whether in cirrhosis-PHT models, ± superimposed inflammation to mimic acute-on-chronic liver failure (ACLF) modulates hepatic nitric oxide synthase trafficking inducer (NOSTRIN) expression, nitric oxide (NO) synthesis, and/or endothelial dysfunction (ED); and (2) Whether the "angiotensin II type 1 receptor blocker" candesartan cilexetil (CC) affects this pathway. CD-1 mice received intraperitoneal carbon tetrachloride injections (CCl4 15% v/v in corn oil, 0.5 mL/kg) twice weekly for 12 wk to induce cirrhosis. After 12 wk, mice were randomized to receive 2-wk oral administration of CC (8 mg/kg) ± LPS. At sacrifice, plasma (biochemical indicators, cytokines, and angiotensin II) and liver tissues (histopathology, Sirius-red stains, and molecular studies) were analysed. Moreover, Nostrin gene knockdown was tested in human umbilical vein endothelial cells (HUVECs). When compared to naïve animals, CCl4-treated animals showed markedly elevated hepatic Nostrin expression (P < 0.0001), while hepatic peNOS expression (measure of eNOS activity) was significantly reduced (P < 0.05). LPS challenge further increased Nostrin and reduced peNOS expression (P < 0.05 for both) in cirrhotic animals. Portal pressure and subsequent hepatic vascular resistance were also increased in all cirrhotic animals following LPS challenge. In CCl4 ± LPS-treated animals, CC treatment significantly reduced Nostrin (P < 0.05) and increased hepatic cGMP (P < 0.01). NOSIP, caveolin-1, NFκB, and iNOS protein expression were significantly increased in CCl4-treated animals (P < 0.05 for all). CC treatment non-significantly lowered NOSIP and caveolin-1 expression while iNOS and NFκB expression was significantly reduced in CCl4 + LPS-treated animals (P < 0.05 for both). Furthermore, Nostrin knockdown significantly improved peNOS expression and associated NO synthesis and reduced inflammation in HUVECs. This study is the first to indicate a potential mechanistic role for the Nostrin-eNOS-NO pathway in cirrhosis and ACLF development. Moreover, this pathway provides a potential therapeutic target given the ameliorative response to Candesartan treatment.

Investigations of an organic-inorganic nanotheranostic hybrid for pancreatic cancer therapy using cancer-in-a-dish andin vivomodels.

The incidence of highly aggressive pancreatic cancer is increasing across the globe and is projected to increase to 18.6% by 2050. The mortality rate for this form of cancer is very high and the 5 y relative survival rate is only about 9%-10%. The 3D pancreatic cancer microenvironment exerts a major influence on the poor survival rate. A key factor is the prevention of the penetration of the chemotherapeutic drugs in the three-dimensional (3D) microenvironment leading to the development of chemoresistance which is a major contributor to the survival rates. Hence,in vitrostudies using 3D cultures represent a better approach to understand the effect of therapeutic formulations on the cancer cells when compared to conventional 2D cultures. In the present study, we have explored three different conditions for the development of a 3D pancreatic tumour spheroid model from MiaPaCa-2 and PanC1 cells cultured for 10 days using Matrigel matrix. This optimized spheroid model was employed to evaluate a multi-functional nanotheranostic system fabricated using chitosan nanoparticles co-encapsulated with the chemotherapeutic agent gemcitabine and gold-capped iron oxide nanoparticles for multimodal imaging. The effect of the single and multiple-dose regimens of the theranostic system on the viability of 3D spheroids formed from the two pancreatic cancer cell lines was studied. It was observed that the 3D tumour spheroids cultured for 10 days exhibited resistance towards free gemcitabine drug, unlike the 2D culture. The administration of the multifunctional nanotheranostic system on alternate days effectively reduced the cancer cell viability after five doses to about 20% when compared with other groups. The repeated doses of the nanotheranostic system were found to be more effective than the single dose. Cell line-based differences in internalization of the carrier was also reflected in their response to the nanocarrier with PanC1 showing better sensitivity to the treatment.In vivostudies revealed that the combination of gemcitabine and magnetic field induced hypothermia produced superior regression in cancer when compared with the chemotherapeutic agent alone by a combination of activating the pro-apoptotic pathway and heat-induced necrosis. Our results reveal that this multi-functional system holds promise to overcome the current challenges to treat pancreatic cancers.

Development and evaluation of a multi-functional organic-inorganic nanotheranostic hybrid for pancreatic cancer therapy.

Pancreatic cancer is a highly invasive disease with low survival rates. The high death rates associated with pancreatic cancer are due to multiple factors including late stage diagnosis, multi-drug resistance, invasive nature and restricted access of the therapeutic moiety to the cancer cells due to the stroma. Smart multifunctional nanocarriers that deliver the therapeutic agent in to the cancer tissue as well as enable imaging of the tissue represent an emerging paradigm in cancer therapy. Accurate and reliable detection of cancerous lesions in pancreas is essential for designing appropriate therapeutic strategy to annihilate the highly aggressive pancreatic cancer. A combination of imaging modalities can enhance the reliability of cancer detection. In this context, we report here a hybrid iron oxide-gold nanoparticle with dual contrast enhancing ability for both magnetic resonance imaging (MRI) and micro-computed tomography (micro-CT) that is co-encapsulated with the nucleotide analogue gemcitabine in a chitosan matrix. The theranostic system displayed enhanced cytotoxicity against PanC-1 pancreatic cancer cells when compared to normal cells over 48 h due to differences in cell internalization. The iron oxide-gold hybrid enabled visualization of the theranostic nanoparticle by MRI as well as micro-CT. Further, the magnetocaloric effect of the iron oxide enabled faster release of the chemotherapeutic agent as well as augmented the cytotoxicity by inducing hyperthermia. This system holds promise for further exploration as an integrated diagnostic and therapeutic platform for pancreatic cancer.

Dataset for compression ignition engine fuelled with corn oil methyl ester biodiesel.

The present data study deals with the experimental analysis of performance, emission and combustion characteristics of CI engine fuelled with corn oil methyl ester biodiesel blend as alternative fuel. A two-step trans esterification process is used to produce the biodiesel. Furthermore, a characteristics study was carried out on the extracted corn oil methyl ester biodiesel blends over conventional fuel. Three different fuel blends namely B10, B20 and B30 were chosen and the performance, emission and combustion characteristics of these are compared to that of conventional diesel fuel. Eddy current dynamometer is used load the engine from no load to maximum load condition. Using AVL DiGAS 444 N gas analyser and AVL 346 smoke meter the emissions and smoke opacity of the fuel blends and diesel were measured respectively. The experimental performance, emission and combustion data's were presented.

Overexpression of bone morphogenetic protein 4 enhances the invasiveness of Smad4-deficient human colorectal cancer cells.

Bone morphogenetic proteins (BMP), a member of the TGF-beta superfamily, have broad activities in regulating various kinds of cellular behaviors. Previously, we have demonstrated that BMP-4 is up-regulated in human colonic adenocarcinoma and promotes the invasive phenotypes of human colorectal cancer HCT116 cells. Smad4 is a central mediator in BMP signaling pathway and it is frequently mutated in metastatic colorectal cancers. To address whether Smad4 was required for enhancing metastatic potentials by BMP-4 in colorectal cancer, we generated BMP-4 overexpressing clones from Smad4-deficient SW480 cells. The growth rate of BMP-4 overexpressing cells was slightly higher than that of empty-vector controls. Overexpression of BMP-4 resulted in an increased expression of vimentin, a marker of epithelial-mesenchymal transition, and caused the changes of cell morphology, spreading and formation of focal adhesions. BMP-4 overexpressing cells increased cell adhesion on fibronectin and collagen, and augmented cell migration and invasion potentials in comparison to empty-vector controls. The induction of cell migration by BMP-4 overexpression was inhibited by BMP-4 siRNA. To further identify the target genes of the elevated BMP-4 signaling in SW480 cells, an oligonucleotide microarray was performed. Among 46,000 genes, 91 genes (65 up-regulated and 26 down-regulated) with 2-fold difference have been identified between BMP-4 overexpressing and empty-vector cells. This study demonstrates that Smad4 is dispensable for enhanced invasiveness of human colorectal cancer cells due to BMP-4 overexpression.

Alteration of Drug Sensitivity in Human Colon Cancer Cells after Exposure to Heat: Implications for Liver Metastasis Therapy using RFA and Chemotherapy.

Radiofrequency ablation (RFA) is gaining popularity for treating colorectal liver metastases by inducing image guided tumor hyperthermia. In order to reduce tumor recurrence, adjuvant therapies have been administered post-RFA. We hypothesized that tumor cells escaping RFA cytotoxicity by being in the sublethal zones of tumor might develop differential behavior toward cytotoxic drugs. Here, we used cultured human colorectal cancer cells to evaluate the interaction between heat treatment and chemotherapeutic agents. Human colon cancer cell lines HT29 and HCT116 were subjected to temperatures of 42 degrees to 50 degrees C for 15 min, in combination with 5-fluorouracil, oxaliplatin, or irinotecan at different sequences. Cytotoxicity was determined by MTT assay. The cell cycle progression was analyzed by flow cytometry with propidium iodide staining. The expression of several genes associated with drug sensitivity was quantitated by real-time RT-PCR before and after heat treatment. Either heat treatment at 45 degrees C by simultaneous or pre-treatment with three different chemotherapeutic agents didn't affect the cytotoxicity of the combined treatment to HT29 and HCT116 cells, except for irinotecan treatment in HCT116 cells. However, when pre-exposure to 45 degrees C, HCT116 cells, but not HT29 cells, developed resistance to these three drugs. In an analysis of cell cycle profile after the drug followed heat treatment, a longer delay in cell cycle progression in HCT116 cells was observed in comparison to HT29 cells. Furthermore, HCT116 and HT29 cells exhibited different expression profiles of several drug-related genes in response to heat treatment at 45 degrees C. An observation of a differential response to the drug and heat treatment sequences between two human colon cancer cell lines suggests that tumor heterogeneity and selection of chemotherapeutic agents need to be under consideration in the clinical setting.

Bone morphogenetic protein-4 inhibits heat-induced apoptosis by modulating MAPK pathways in human colon cancer HCT116 cells.

Cancer thermotherapy and radiofrequency ablation (RFA) have been adopted as modalities for treating various kinds of cancer. We have previously demonstrated that bone morphogenetic protein-4 (BMP-4) is up-regulated in colonic adenocarcinoma. Here, we investigated whether an increase of BMP-4 expression changes cellular response to heat treatment in human colon cancer HCT116 cells. BMP-4 overexpressing HCT116 cells generated by stable transfection showed a significantly increased survival rate and a decreased apoptotic rate in comparison to empty vector controls after heat treatment at 45 degrees C for 20min. The expression levels and pattern of HSP90, HSP70, and HSP27 after heat treatment were similar between these two cell lines. There was no difference in expression levels of Bcl-2 and Bax in these two cell lines and their expression remained unchanged after heat treatment. Both activities of the extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) were stimulated by heat in these cells. Comparatively, BMP-4 overexpressing cells had an intense and prolonged ERK activation, while a less intense and short JNK activation. Correspondingly, treatment of BMP-4 overexpressing cells with noggin, a BMP-4 antagonist, resulted in a reduction of heat-activated ERK but an increase of heat-activated JNK and significantly increased heat-induced apoptotic rate. These results indicate that BMP-4 can protect colon cancer cells from heat-induced apoptosis through enhancing the activation of ERK as well as inhibiting the activation of JNK.

Bone morphogenetic protein-4 is overexpressed in colonic adenocarcinomas and promotes migration and invasion of HCT116 cells.

Bone morphogenetic protein (BMP), a member of the TGF-beta superfamily, is involved in development, morphogenesis, cell proliferation and apoptosis. Dysregulation of BMP signaling has been suggested in tumorigenesis. In an analysis of human colon normal mucosa and tumors at different stages by immunohistochemistry, we observed that the intensity of BMP-4 staining in late-adenocarcinomas was stronger than that in normal mucosa and adenomas, while there was no difference in the staining of its receptors (BMPR-IA and BMPR-II) at all stages. The up-regulation of BMP-4 was further validated in another panel of tumor tissues by real-time RT-PCR, showing that BMP-4 mRNA levels in primary colonic carcinomas with liver metastasis were significantly higher than that in the matched normal mucosa. In order to understand the functional relevance of BMP-4 expression in colon cancer progression, BMP-4-overexpressing cell clones were generated from HCT116 cells. Overexpression of BMP-4 did not affect the HCT116 cell growth. The cells overexpressing BMP-4 became resistant to serum-starvation-induced apoptosis and exhibited enhanced migration and invasion characteristics. Overexpression of BMP-4 changed cell morphology to invasive spindle phenotype and induced the expression and activity of urokinase plasminogen activator (uPA). These results indicate that BMP-4 confers invasive phenotype during progression of colon cancer.

Induction of p21WAF1 expression protects HT29 colon cancer cells from apoptosis induced by cryoinjury.

BACKGROUND: Cryotherapy is a method of in situ destruction of tumors by freeze/thaw mechanisms. Cancer cells located in the peripheral zone of the tumor undergoing cryotherapy can die by apoptosis. We hypothesized that p21WAF1 is involved in the mediation of cryotherapy-induced apoptosis. METHODS: HT29 cells grown on a plate were subjected to -10 degrees C and returned to 37 degrees C for various periods of time. Cells were analyzed by flow cytometry, Western blot, and reverse transcriptase-polymerase chain reaction for determining cell-cycle distribution, p21WAF1 protein expression, and messenger RNA levels, respectively. The p21WAF1 expression in nude mouse tumor xenografts after cryotherapy was examined by immunofluorescence staining. A series of the p21WAF1 promoter cloned into a luciferase reporter vector were transfected into HT29 cells for identifying the response element to cryoinjury. Antisense oligodeoxynucleotide (ODN) was applied to examine the effect of p21WAF1 expression on cryotherapy-induced apoptosis. RESULTS: Both protein and messenger RNA of p21WAF1 were induced by cryoinjury in cultured cells and tumor xenografts. Deletion analysis of the p21WAF1 promoter revealed that a region from -121 to -95 base pairs was responsible for the activation and that this activation was p53 independent. HT29 cells arrested at the G1 phase after cryoinjury. The cryotherapy-induced apoptotic rate in HT29 cells was increased in the presence of antisense p21WAF1 ODN in comparison to the random ODN. CONCLUSIONS: Induction of p21WAF1 increases tumor cell survival and may result in recurrences at treated sites after cryotherapy. Combining antisense ODN targeted against p21WAF1 and cryotherapy may improve clinical outcomes in the treatment of colorectal cancer.

Heat shock protein 70 is induced in mouse human colon tumor xenografts after sublethal radiofrequency ablation.

BACKGROUND: Radiofrequency ablation (RFA) destroys tumor cells by generating high temperatures through ionic vibration. Tumor recurrence may be a direct function of sublethal temperature. Further, a set of proteins called heat shock proteins (HSPs) can be synthesized under heat stress to facilitate recovery of tumor cells from heat damage. METHODS: Subcutaneous xenografts were induced in nude mice by injection with HT29 human colon cancer cells. The tumors were exposed surgically and subjected to RFA. The tumors were randomly assigned to achieve a target tumor temperature of 42 degrees C, 45 degrees C, or 50 degrees C. Total RNA and cell lysates were isolated from tumor tissues and subjected to reverse transcription-polymerase chain reaction and Western blot analyses, respectively, at various time points after treatments for assessing HSP expression. For in vitro experiments, HT29 cells were subjected to variable temperatures, and HSP expression was assayed. RESULTS: During a 50-day follow-up, the recurrence rates were 0% at 50 degrees C, 30% at 45 degrees C, and 100% at 42 degrees C. The messenger RNA and protein levels of HSP90 and HSP27 remained unchanged after RFA at 45 degrees C; however, HSP70 was induced at 4 and 10 hours after RFA. In vitro HT29 culture cells subjected to a heated water bath exhibited a cellular sensitivity to heat and change of HSP expression similar to those in tumor xenografts subjected to RFA. CONCLUSIONS: Our data establish the requisite heat parameters during RFA for human colon tumors in vitro and in vivo. Because HSP70 plays an important role in protecting cell death from a variety of stresses, HSP70 could be a potential target for enhancing the efficacy of RFA.

Development of cross-resistance between heat and cisplatin or hydroxyurea treatments in FaDu squamous carcinoma cells.

BACKGROUND: Induction of hyperthermia by radiofrequency ablation is gaining popularity in treating a variety of solid tumors. This study examined an impact of sublethal heat treatment interacted with chemotherapeutic drugs on the survival of head and neck squamous carcinoma cells using in vitro model. MATERIALS AND METHODS: FaDu cells were subjected to heat treatment at 42 degrees C or 45 degrees C for 15 min either before or after exposure to cisplatin or hydroxyurea. The survival of cells was determined by 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide assay. The RNA and protein levels of various heat shock proteins were examined by reverse transcription polymerase chain reaction and Western blot analysis, respectively. Cell cycle progression was analyzed by flow cytometry with propidium iodide staining. RESULTS: FaDu cells preheated to 45 degrees C exhibited an increased resistance to hydroxyurea but not to cisplatin. The heat treatment resulted in induction of HSP70 expression at transcript and protein levels, but there was no change in expression of HSP90beta and HSP27. After heat treatment, cells accumulated in S-phase at 3 h and proceeded to G(2)/M phase at 24 h. When cells pre-exposed to drugs for 24 h, the cisplatin-treated cells exhibited a higher thermotolerance than the hydroxyurea-treated cells at heat treatment of 45 degrees C. Cisplatin and hydroxyurea caused cells to accumulate in S-phase and increased the protein expression of HSP27 but not HSP90beta and HSP70. CONCLUSION: FaDu cells surviving the heat treatment expressed HSP70 and disrupted cell cycle progression, which resulted in developing a resistance to subsequent hydroxyurea treatment. However, the heat treatment did not have an effect on the sensitivity to cisplatin. In the reversed procedure, pre-exposure to hydroxyurea and cisplatin resulted in developing a thermotolerance.

Apoptosis induced by cryo-injury in human colorectal cancer cells is associated with mitochondrial dysfunction.

Cryotherapy, a method of in situ ablation, is used in the treatment of colorectal liver metastases with variable results. During the treatment, the central area of treated tumor undergoes necrotic destruction by lethal cryo-injury; however, the cellular response of tumor exposed to sublethal cryo-injury at the peripheral zone is unclear. In our study, we have identified the induction of apoptosis by cryo-injury at -10 degrees C in 4 colorectal cancer cell lines (HT29, HCT116, KM12C and KM12SM). The apoptosis was characterized by chromatin condensation, transferase-mediated dUTP nick end-labeling (TUNEL) staining, proteolytic cleavage of poly(ADP-ribose) polymerase (PARP) and cytokeratin 18, and activation of caspase-3. The occurrence and intensity of cryo-induced apoptosis did not correlate with the functional status of p53 in the cell lines studied. The expression of anti-apoptotic proteins (Bcl-2, Bcl-X(L)) and pro-apoptotic proteins (Bax, Bcl-X(S), Bad, and Bak) in response to cryo-injury varied in this cell line panel. The basal level of Bcl-2/Bax protein ratio correlated inversely to the apoptotic rate. We further demonstrated that Bax level decreased in cytosol and increased in mitochondria, followed by a loss of mitochondrial membrane potential after cryo-injury in HT29 cells. These findings indicate that cryo-injury induces apoptosis in colorectal cancer cells via disruption of mitochondrial integrity. The cryo-induced apoptosis was also identified in a nude mouse tumor xenograft model. Our elucidation of the apoptosis pathway induced by cryo-injury implies that synergistic combination of cryosurgery with pharmacological agents that augment of apoptosis induction may have clinical relevance in treating colorectal liver metastasis.

Resection of liver metastases: state of the art.

In this article, we present current surgical perspectives on the management of liver metastases, with a focus on state-of-the-art resection, by drawing on clinical data provided in the medical literature. Metastases from colorectal cancer are the most amenable to resection, when considering long-term benefits. To highlight the evidence-based rationalefor surgical resection, a succinct comparison of all treatment modalities for liver metastasis from colorectal cancer is provided. Special emphasis has been placed on the current status of perioperative patient evaluation, durability of results, prognosticators, technical advances, and alternative treatment strategies. In various areas, we have attempted to delineate any improvements on the horizon. The role of surgical resection in noncolorectal metastases is also discussed.

Induction of apoptosis in human colon carcinoma cells HT29 by sublethal cryo-injury: mediation by cytochrome c release.

Cryosurgery is an emerging treatment for human solid tumors, notably colorectal liver metastasis. Cryosurgical procedures generate a thermal gradient of from at least -50 degrees C at the center of the tumor being treated to about 0 degrees C at the periphery. Cell death occurs by necrosis in the center, while the peripheral zone of frozen tumor harbors a mix of viable and dead tissue. In order to understand the mechanisms of cell death and survival in this peripheral area at risk for tumor recurrence, we have established an in vitro freezing system that mimics in vivo conditions of sublethal injury. HT29 colon cancer cells were subjected to freezing temperatures from -6 degrees C to -36 degrees C, thawed at room temperature for 30 min and rewarmed at 37 degrees C for a period of time. Post-freeze-thaw, cryolytic cells were evaluated by trypan blue exclusive assay. We also identified apoptotic cells after rewarming by cell shrinkage, nucleic condensation, TUNEL assay, DNA fragmentation and PARP degradation. The intensity of cryolysis and apoptosis was increased by lowering the freezing temperature. At -36 degrees C, all cells were dead immediately after freeze-thaw. A kinetic analysis of cryo-induced apoptosis showed that the commitment to enter apoptosis occurred right after the freeze-thaw period and lasted less than 8 hr after rewarming. We further demonstrated that freezing triggers one of the caspase cascade involved in apoptosis: release of cytochrome c from mitochondria to cytosol, followed by activation of caspase-9 and degradation of PARP. These results indicate the death of cancer cells under cryo-treatment at sublethal freezing temperature can be attributed 2 different modes, cryolysis as well as apoptosis. HT29 cells carrying p53 mutant have very quick response for induction of apoptosis by cryo-treatment and contain an intact pathway of caspase cascade. Further studies will address if mechanisms in cells with wild-type p53 will differ.

Total pelvic exenteration and reconstruction for locally invasive recurrent sarcoma of the perineum.

Extensive primary tumors and locally recurrent tumors of the pelvis or perineum are difficult to manage. We describe the techniques necessary to perform total pelvic exenteration with en bloc resection of the perineum and genitalia for treatment of recurrent sarcoma of the perineum. Wide excision of the sarcoma with negative margins can be achieved by resecting the inferior portion of the pubic symphysis. An absorbable mesh sling may be used to suspend the small bowel above the pelvis, facilitating postoperative radiation. A catheterizable continent urinary reservoir avoids the necessity of two stomas and improves quality of life. Adequate tissue coverage can be attained by myocutaneous gracilis flaps that promote wound healing.

In vitro activation of irinotecan to SN-38 by human liver and intestine.

BACKGROUND: Irinotecan (CPT-11) is hydrolyzed by carboxyl esterase to the active metabolite SN-38 and oral irinotecan could undergo intestinal and hepatic activation. MATERNALS AND METHODS: Irinotecan was incubated with S9 fractions of human liver and intestinal tissues and the specific activity was determined based on the formation rate of SN-38. RESULTS: Irinotecan was hydrolyzed to SN-38 by hepatic and intestinal S9 fractions with mean (+/- SD) specific activities (pmoles/min/mg) of: liver (8.57 +/- 10.4, n = 8), duodenum (5.06 +/- 3.7, n = 4), jejunum (6.44 +/- 2.8, n = 5), ileum (4.81 +/- 2.4, n = 5), colon (1.93 +/- 1.5, n = 6) and rectum (0.82, n = 1). When incubated with S9 fractions obtained from tumor tissues, there appeared to be a decrease in SN-38 formation compared to matched normal liver and colon tissues. CONCLUSION: Irinotecan undergoes conversion to its active metabolite in human intestinal S9 fractions and there is variability in the extent of SN-38 formation. The localized intestinal activation of irinotecan to SN-38 may provide a rationale for the development of oral irinotecan for gastrointestinal malignancies but could also cause mucosal damage leading to toxicity.

Minimally invasive staging of patients with melanoma: sentinel lymphadenectomy and detection of the melanoma-specific proteins MART-1 and tyrosinase by reverse transcriptase polymerase chain reaction.

BACKGROUND: A minimally invasive standard has yet to be developed for sentinel lymphadenectomy, and many patients undergo this procedure in the main operating room under general anesthesia. These patients often have microscopic metastases in sentinel nodes that could be missed by histopathologic examination. Techniques of reverse transcriptase polymerase chain reaction (RT-PCR) could detect these metastases if the nodes could be preserved intraoperatively. STUDY DESIGN: Fifty patients with melanoma > or = mm thick underwent sentinel lymphadenectomy under local anesthesia in an outpatient surgical unit. Sentinel nodes were identified using blue dye and technetium-99 sulfur colloid and a hand-held gamma probe. Each node was sectioned, with half sent for routine histopathologic study and half preserved in liquid nitrogen. We used RT-PCR to detect mRNA for tyrosinase and Melanoma Antigen Recognized by T cells-1 (MART-1). RESULTS: All patients were able to tolerate sentinel lymph node biopsy under local anesthesia. Sentinel lymph nodes were obtained in 100% of our patients, and usable mRNA was harvested from all but five. Ten patients had positive sentinel node(s) by standard histopathologic examination, and all of these nodes were also positive for MART-1 and tyrosinase. Three patients with negative results by histopathology had positive results by RT-PCR analysis. The average cost of these outpatient operations was 38% less than the same operations performed in the main operating room under general anesthesia. CONCLUSIONS: Sentinel lymphadenectomy under local anesthesia in an outpatient setting and intraoperative lymph node preservation in liquid nitrogen are both feasible. Both tyrosinase and MART-1 are promising markers in the detection of occult melanoma in lymph nodes.

Experimental basis for hepatic cryotherapy.

Hepatic cryotherapy has emerged as a viable therapeutic option in the treatment of liver tumors, both primary and secondary. We will provide the experimental basis for its application in the liver with particular reference to safety and efficacy issues. Also discussed are experiments with the use of hepatic cryotherapy in animals that have demonstrated the effectiveness of the technique, and improvements in technology that have enabled better delivery. The future of this modality looks bright with increasing advances in technology to enable better application. Its use in combination with other treatment modalities could offer improved success in treating liver tumors.

Chemotherapy for metastatic melanoma during pregnancy.

Melanoma during pregnancy represents a difficult problem. Although surgery is a definitive therapy for early-stage disease, rapidly progressive metastatic disease in pregnancy requires a series of more difficult decisions. We report the use of combination chemotherapy with tamoxifen, carmustine, dacarbazine, and cisplatin in a patient with metastatic melanoma during the second trimester of pregnancy. The patient received 2 cycles of chemotherapy prior to delivery of a healthy 1520-g baby girl at 30 weeks gestation. Placental pathology, however, revealed malignant melanoma in the intervillous space and melanin pigment granules in villous Hofbauer cells and synctial trophoblasts. This report also reviews the literature, assessing the importance of pregnancy as a prognostic factor, the effects of metastasis on the products of conception, and the use of chemotherapy in pregnancy. We conclude that combination chemotherapy can be administered in the second trimester of pregnancy for the treatment of rapidly progressive melanoma without interfering with the successful maturation and delivery of an infant of 30 weeks gestation.