A Cost Analysis of Healthcare Episodes Including Day-Case Bariatric Surgery (Roux-en-Y Gastric Bypass and Sleeve Gastrectomy) Versus Inpatient Surgery.

BACKGROUND: Assessing the medico-economic outcomes of a healthcare pathway including day-case bariatric surgery versus the conventional pathway. METHODS: This economical evaluation is a prospective cohort study with historical controls. Between March 2019 and December 2020, 30 patients eligible for bariatric surgery were considered in the day-case group. Surgical procedures included sleeve gastrectomy and Roux-en-Y gastric bypass. The day-case pathway included patient education, post-discharge follow-up by a community nurse twice-daily and standardized communications to surgeons. Day-case patients were paired with 30 inpatients, based on the type of intervention, age, and ASA status. The primary outcome was the cost of care episodes from the preoperative visit to the 30-day postoperative visit. Micro-costing methodology and activity-based costing were used. Secondary outcomes included length of hospital stay, rate of unanticipated events, and patient' satisfaction assessment. RESULTS: Male-to-female ratio was 1/2. In the day-case versus inpatient group, age, number of associated medical conditions, and BMI (42.9 ± 4.9 versus 42.6 ± 4.6, p > 0.05) were similar. In the day-case group, there were 7 overnight stays (23.3%), 3 readmissions (10%), and 4 unscheduled consultations (13.3%). The overall length of hospital stay was significantly shorter (0.65 ± 0.33, versus 2.9 ± 0.4 days, p < 0.0001). The complication rate was 6.6% in both groups. The cost of the care episode was € 4272.9 ± 589.7 for the day-case group versus € 4993.7 ± 695.6 for inpatients, corresponding to a 14.4% cost reduction (p = 0.0254). CONCLUSIONS: Day-case bariatric surgery appears to be safe and beneficial in terms of costs. It involves a specific organization with postdischarge follow-up. TRIAL REGISTRATION: ClinicalTrial.gov: NCT04423575.

18F-Fluorocholine PET and 4D-CT in Patients with Persistent and Recurrent Primary Hyperparathyroidism.

Patients with primary hyperparathyroidism (pHPT) can develop persistent (P-pHPT) or recurrent (R-pHPT) disease after parathyroidectomy. Before recommending reoperation, recurrence must be accurately identified because of the high risk of complications. Our study evaluates 18F-fluorocholine (18F-FCH) PET/CT and 4D-CT integrated in PET/4D-CT in patients with P-pHPT/R-pHPT. Patients with P-pHPT/R-pHPT investigated by 18F-FCH PET/4D-CT between May 2018 and March 2021 were retrospectively included. Forty-two patients were included, 37 of whom underwent 4D-CT. The sensitivity and detection rate (DR%) were 95% and 88% for 18F-FCH PET/CT and 70% and 63% for 4D-CT, respectively. PET/CT and 4D-CT were concordant in 18/24 glands and concordant and positive in 15/24 (63%) glands. Discordant results were obtained for 6/24 glands. The surgical success rate was 65%. PET/CT showed significantly higher sensitivity than 4D-CT. Dynamic CT allowed the identification of no additional glands missed by PET/CT, and the combination of the 2 techniques did not improve the sensitivity or DR%. 18F-FCH PET/CT appears to be a valuable technique to accurately detect hyperfunctioning parathyroid tissue in patients with P-pHPT/R-pHPT and is better than 4D-CT. Except for cases with doubtful locations of PET targets that may require 4D-CT for surgical guidance, standard nonenhanced 18F-FCH PET/CT can be effectively recommended in patients with P-pHPT/R-pHPT before reoperation.

Dual-Time-Point 18F-Fluorocholine PET/CT Improves Characterization of Thyroid Nodules in Patients Referred for Primary Hyperparathyroidism: A Proof of Concept Study.

PURPOSE: Thyroid nodules frequently coexist with primary hyperparathyroidism (pHPT). Because of the increasing use of 18F-fluorocholine (18F-FCH) PET/CT in patients with pHPT, evaluation of its clinical utility for thyroid nodules characterization in this population is of paramount importance. Herein, we investigate the value of dual-point 18F-FCH PET/CT in the diagnosis of thyroid cancer in patients referred for pHPT imaging who have thyroid nodules. PATIENTS AND METHODS: All pHPT patients who underwent a dual-time point 18F-FCH PET/CT (at 5 and 60 minutes postinjection) between July 2019 and December 2020 were analyzed. Only those with a thyroid nodule greater than 10-mm and pathological analysis (criterion standard) were included. Nodule-to-thyroid SUVmax ratio was calculated at the 2 study points, as well as the 18F-FCH washout index (WO%). RESULTS: Twenty-seven patients (32 nodules) were included in this study. The final diagnoses were as follows: 27 benign nodules including 2 NIFTPs (noninvasive follicular thyroid neoplasm with papillary-like nuclear features) and 5 cancers of follicular origin. Early uptake ratio was significantly higher in malignant lesions than in benign nodules (P = 0.0008). Thyroid cancers were also characterized by a marked 18F-FCH washout index (WO% benign vs cancer: 2.9% ± 4.1% vs 45.5% ± 13.4%, P = 0.0001). Using a WO% threshold of 22.1%, 25/27 benign nodules and 5/5 malignant lesions were accurately classified (sensitivity of 100%, specificity of 92.6%, positive predictive value of 71.4%, and negative predictive value of 100%). The false-positive findings were related to the 2 NIFTPs that share similarities with thyroid cancer. CONCLUSIONS: Our preliminary results suggest to perform a dual-time-point PET/CT acquisition protocol in pHPT patients with uncharacterized centimeter thyroid nodules. However, the real impact of these promising results should be assessed by prospective studies on a larger cohort of patients.

Indications and Long-Term Outcomes of Conversion of Sleeve Gastrectomy to Roux-en-Y Gastric Bypass.

PURPOSE: Long-term results on sleeve gastrectomy (SG) with more than 10 years report patients needing sleeve revision for weight loss failure, de novo gastroesophageal reflux (GERD), or sleeve complications. The aim of this study was to analyze the results of laparoscopic conversion of failed SG to Roux-en-Y gastric bypass (RYGB). MATERIALS AND METHODS: Retrospective review of a prospectively institutional maintained database to identify patients who underwent conversion of SG to RYGB between 2012 and June 2020. RESULTS: Sixty patients(50 females) underwent conversion to RYGB. Average time to conversion was 5.6 years (2-11). Mean %WL and TWL after SG were respectively 26±8.8% and 33.2±14.1kg. Mean BMI at the time of RYGB was 38.1±7.1 kg/m2. Mean follow-up was 30.4±16.8 months (6-84). Available patients at each time of follow-up: 1 year 59 (98.3%); 2 years 47 (78.3%); 3 years 39 (71.6%); and 5 years 33 (55%). Patients were divided according to indication for revision in weight regain/insufficient weight loss (30 patients) group 1 and GERD/complications (25 patients) group 2. Percentage of excess weight loss at 1, 3, and 5 years follow-up after bypass was for group 1 40.3±17.6, 34.3±19.5, and 23.2±19.4 and for group 2 90.4±37, 62.6±28.2, and 56±35.02. Total weight loss at last follow-up since sleeve was respectively 31kg in group 1 and 46.7kg in group 2 (p=0.002). No mortality was observed. Thirty-day complication rate was 3.3%. CONCLUSION: RYGB after SG is a safe and effective revisional procedure to manage weight regain and de novo GERD, to address complications, and to improve comorbidities.

Original Preoperative Localization Technique of Parathyroid Adenomas by 3-Dimensional Virtual Neck Exploration.

Objective. Preoperative imaging in primary hyperparathyroidism (PHPT) is essential for planning of parathyroidectomy-particularly for selection of a minimally invasive approach. The objective of this cohort study was to evaluate the diagnostic precision of 3D virtual neck exploration (3D-VNE), to evaluate its impact on choice of surgical approach, and to document the correlation with long-term outcomes. Methods. 235 consecutive patients with PHPT were studied (January 2014 to December 2018), with 6-month follow-up. 220 patients had a preoperative computed tomography (CT), 172 of these had a 3D-VNE based on the CT, and 226 patients had a Tc-99m sestamibi scan. Results. Sensitivity of exact, per gland, adenoma localization was 57.09% (95% CI: 50.85-63.10%) for nonspecialized radiologist interpretation of CT scan, 58.17% (95% CI: 51.99-64.10%) for Tc-99m sestamibi scan, and 90.21% (95% CI: 85.21-93.64%) for 3D-VNE, and thereby favoring 3D-VNE compared to CT scan alone (OR 34.5, 95% CI: 9.19-290.56%, P < 2.2 × 10-16) and to Tc-99m sestamibi scan (OR 16.25, 95% CI: 6.05-61.42%, P = 3.1 × 10-15). Specificity was 87.38% for CT scan, 86.36% for 3D-VNE, and 90% for Tc-99m sestamibi scan (P > .05). The cure rate was 100%. The long-term recurrence rate (RR) was 2.978%. The RR was 1.324% in the video-assisted parathyroidectomy group of 151 patients and 5.952% in the group of 84 patients with cervicotomy (P = .0459). Conclusion. CT-based 3D-VNE proved to be the most accurate localizing study in PHPT and aided in selecting patients for targeted minimally invasive parathyroidectomy, which was associated with the lower recurrence rate. 3D-VNE could be proposed as a first-line imaging study in patients with PHPT.

18F-Fluorocholine PET and Multiphase CT Integrated in Dual Modality PET/4D-CT for Preoperative Evaluation of Primary Hyperparathyroidism.

The present retrospective study evaluates the diagnostic value of integrated 18F-Fluorocholine positron emission tomography/four-dimensional contrast-enhanced computed tomography (18F-FCH PET/4D-CT) as second-line imaging in preoperative work-up of primary hyperparathyroidism (pHPT), and compares 18F-FCH PET with 4D-CT. Patients with pHPT and negative/discordant first-line imaging addressed for integrated 18F-FCH PET/4D-CT were retrospectively selected. Sensitivity and detection rate (DR%) of 18F-FCH PET/CT, 4D-CT, and PET/4D-CT were calculated according to the per patient and per lesion analyses, and afterwards compared. Histology associated with a decrease more than 50% of perioperative parathyroid hormone (PTH) blood level was used as a gold standard. Persistent high serum PTH and calcium levels during a 6-month follow-up was considered as presence of pHPT in both operated and non-operated patients. 50 patients (55 glands) were included. 44/50 patients (88%) were surgically treated. On a per patient analysis, sensitivity was 93%, 80%, and 95%, and DR% was 82%, 68%, and 84%, respectively for PET/CT, 4D-CT, and PET/4D-CT. PET/CT was more sensitive than 4D-CT (p = 0.046). PET/4D-CT performed better than 4D-CT (p = 0.013) but was equivalent to PET/CT alone. On a per gland analysis, sensitivity PET/CT, 4D-CT, and PET/4D-CT was 88%, 66%, and 92%, and DR% was 79%, 57%, and 83%, respectively. PET/CT and PET/4D-CT were more sensitive than 4D-CT alone (p = 0.01, p < 0.001, respectively). However, PET/CT and PET/4D-CT performed similarly. In conclusion, 18F-FCH PET provides better identification of hyperfunctioning parathyroids than 4D-CT and the combination of both did not significantly improve diagnostic sensitivity. Further investigations involving larger populations are necessary to define the role of 18F-FCH PET/4D-CT as a "one-stop shop" second-line imaging in preoperative work-up of pHPT, especially considering the additional patient radiation exposure due to multi-phase CT.

Postoperative Remote Monitoring with a Transcutaneous Biosensing Patch: Preliminary Evaluation of Data Collection.

Introduction. Connected systems transmitting vital parameters could well represent a tool to shorten postoperative hospital stay while providing continuous remote patient monitoring and potentially detect the onset of complications. Our aim was to analyze the functionality of a transcutaneous biosensing data collection patch in morbidly obese patients. Materials and Methods. An adhesive patch (The HealthPatch MD™) was applied to patients' chests postoperatively. The patch was connected to a tablet via a bluetooth network to collect the heart rate, respiratory rate, skin temperature, and posture recognition data. The tablet conveyed data to a secure health data central server by means of a WiFi or 3G/4G transmission. Data were stored in a digital health platform to which health care professionals could connect. The evaluation focused on the volume, quality, and security of data transmission. A pilot phase involved 10 patients. Thirty-three additional patients undergoing bariatric surgery were included in the experimental phase. Results. The mean length of stay was 2.28 days (range: 2-5 days). The mean time of patch application was 51 ± 25.2 hours per patient (range: 19-139 hours), totalizing 1,683 hours of recording for the 33 patients included. During this time, a total of 7.562.531 data measurement points were collected and transmitted to the e-health platform via the patch. Two total disconnections and two partial disconnections were observed. The acquisition of patient postural data was unreliable. Conclusions. Connected telemetry for remote postoperative monitoring is promising. However, it is still limited by data transmission problems.

Intraoperative Probe-Based Confocal Endomicroscopy to Histologically Differentiate Thyroid From Parathyroid Tissue Before Resection.

BACKGROUND: Frozen section is the standard method to histologically distinguish parathyroid tissue from thyroid tissue during endocrine neck surgery. Frozen section can be time-consuming and costly. Its drawback is that it is to be performed only after the removal of a suspected pathological tissue. This study demonstrates the use of probe-based confocal laser endomicroscopy (pCLE) to confirm histology prior to tissue resection. DESIGN: A prospective, single-institution, nonrandomized study was conducted. No sample size calculation was performed for this observational trial. The primary objective was the description of histological rendering of normal and pathological tissues through pCLE. Real-time in vivo fluorescence microscopy imaging was performed with the CystoFlex UHD probe after intravenous injection of 2.5 mL of 10% fluorescein sodium. RESULTS: Eleven patients with hyperparathyroidism and thyroid conditions were included. A total of 104 videos showing thyroid, parathyroid, adipose tissue, muscle, laryngeal nerve, and lymph nodes were recorded. Videos were compared with visual information and pathological samples (when sampling was indicated). Thyroid tissue could be identified based on the presence of colloid follicles (intensely fluorescent area surrounded by a small ridge of low-fluorescence epithelial cells) including the pathognomonic aspect of resorption vacuole. Parathyroid tissue could be identified based on a regular, "diamond-shaped" capillary network encompassing parathyroid chief cells. Blinded reinterpretation of pCLE videos demonstrated an 89.3% sensitivity and a 90% specificity as compared with histology in tissue recognition. CONCLUSION: This pilot study describes representative renderings of intraoperative pCLE to nontraumatically differentiate thyroid, parathyroid, and lymph nodes before surgical removal.

Prospective Trial on Probe-Based Confocal Laser Endomicroscopy for the Identification of the Distal Limit in Rectal Adenocarcinoma.

BACKGROUND: Intraoperative decision of the level of distal resection in rectal cancer is often imprecise, based exclusively on digital examination and pretherapeutic imaging. DESIGN: Prospective, single institution, nonrandomized trial ( ClinicalTrial.gov identification no. NCT01887509) to evaluate the contribution of probe-based confocal laser endomicroscopy (pCLE) to establish the optimal resection margin of rectal adenocarcinoma. The primary outcome was the concordance in the identification of lower tumor margins between pCLE and histopathology. For each patient, pCLE examination was performed on nonneoplastic and neoplastic aspects of the distal tumor margin, before and after neoadjuvant chemoradiation, or preceding surgery, if chemoradiation was not required. Biopsies were taken at the same locations. The intraclass correlation coefficient was determined. RESULTS: Twenty-one patients were enrolled. Thirteen patients completed the full study. Six patients completed imaging only before chemoradiation. Two patients retracted their consent after inclusion. A total of 134 videos and corresponding histopathology samplings were analyzed. The sensitivity and specificity of in vivo pCLE interpretation were 0.915 (95% confidence interval [CI] = 0.840-0.970) and 0.736 (95% CI = 0.657-0.821), respectively. The sensitivity and specificity of the blinded pCLE reinterpretation were 0.930 (95% CI = 0.858-0.980) and 0.688 (95% CI = 0.600-0.770), respectively. No deep layer tumor infiltration was encountered in the samplings with superficial healthy layers. The intraclass correlation coefficient for in vivo pCLE interpretation and blinded pCLE reinterpretation were 0.747 (95% CI = 0.257-0.993) and 0.766 (95% CI = 0.280-0.995), respectively. CONCLUSIONS: This supports the concordance between pCLE and histopathology in identifying the "tumor-free" limit of a rectal tumor preceding resection.

Feasibility and reliability of pancreatic cancer staging using a new confocal non-fluorescent microscopy probe: a double-blind study in rats.

BACKGROUND: Surgical management of pancreatic cancer depends on tumor resectability and staging. Lymph node (LN) metastases represent an important decision-making factor when it comes to surgical treatment. AIMS: To evaluate a new in vivo, endoscopic confocal microscopy (CM) system not requiring fluorescence markers, for detection and staging of pancreatic cancer in rats. METHODS: A confocal system consisting of a confocal scanning laser operating in reflection mode and a dedicated rigid Hopkins rod-lens endoscope were used for in vivo imaging in a rat model of pancreatic ductal adenocarcinoma. A double-blind study compared CM to standard histology in (1) the detection of tumors in rat bearing cancer (n = 11) and controls (n = 6), and (2) in the detection of local nodal involvement at 3 and 6 weeks after tumor induction. RESULTS: CM detected all pancreatic tumors with 100 % sensitivity and specificity and identified 15 metastatic LNs with an average adenocarcinoma nodule diameter of 2.3 mm (range from 1 to 4.2 mm) out of the 66 examined. CM demonstrated a sensitivity of 87.5 % and a specificity of 98 % in LN detection. The Spearman's rank correlation/rho calculator was of 0.87. CM demonstrated a negative predictive value of 96.1 % and a positive predictive value of 93.3 % in the detection of metastatic LNs. CONCLUSIONS: Interpretation of confocal images has a high concurrence rate with histopathology examination for primary tumor and lymphatic involvement detection making it a promising technique for in vivo real-time detection and staging of pancreatic cancer. Larger studies are warranted to confirm these preliminary results.

Survival variability of controls and definition of imaging endpoints for longitudinal follow-up of pancreatic ductal adenocarcinoma in rats.

BACKGROUND: The 3Rs guideline is the gold standard for ethics in animal experimentation. Two of those rules, namely refinement and reduction, require further improvement. The objective of this study was to define pathways to better compliance with these prerequisites. Two methods which move us in this direction are: (1) using small animal imaging techniques for pancreatic ductal adenocarcinoma (PDAC) follow-up and (2) reduction of the number of control animals included in a study of PDAC progression under treatment. MATERIALS AND METHODS: Firstly, we used MicroCT scan to diagnose events showing PDAC progression prior to any clinical symptoms to thereby define more humane endpoints identifiable before any painful phenomenon is observed. Secondly, in order to test the hypothesis of using a reference control group in all preclinical studies of a new treatment of PDAC, we investigated the stability of the results obtained with the control groups in three successive identical studies comparing placebo and gemcitabine in tumor-bearing Lewis rats. RESULTS: Two imaging endpoints were found. The first was the observation of a liver metastasis assessing PDAC diffusion and, earlier than liver metastasis, the presence of bands of fluid along the flanks, with more or less a medial displacement of bowel and solid viscera, reflecting a peritoneal ascites. Results of the longitudinal follow-up of rats in the gemcitabine study revealed heterogeneity in the survival rate in the three control groups, as opposed to the survival rate in the three treated groups which did not differ statistically. As a result, the significance of improved survival with chemotherapy varied greatly according to the control group used for the comparison, ranging from no impact to a highly significant effect. CONCLUSION: The early detection by the means of animal imaging of one or more signs indicating the onset of a critical step in the development of the disease (e.g., ascites or/and metastasis) allows the researcher to prevent the occurrence of animal pain, thereby ensuring better animal welfare. However, using a single standard control group in an effort to use fewer animals for a given model runs such a significant risk of false results that it mars the entire study. Although reducing the number of animals in a study remains the gold standard of our experimental practice, in this case it would come at the price of a loss of validity of the results.

Multimodal imaging of a humanized orthotopic model of hepatocellular carcinoma in immunodeficient mice.

The development of multimodal strategies for the treatment of hepatocellular carcinoma requires tractable animal models allowing for advanced in vivo imaging. Here, we characterize an orthotopic hepatocellular carcinoma model based on the injection of luciferase-expressing human hepatoma Huh-7 (Huh-7-Luc) cells in immunodeficient mice. Luciferase allows for an easy repeated monitoring of tumor growth by in vivo bioluminescence. The intrahepatic injection was more efficient than intrasplenic or intraportal injection in terms of survival, rate of orthotopic engraftment, and easiness. A positive correlation between luciferase activity and tumor size, evaluated by Magnetic Resonance Imaging, allowed to define the endpoint value for animal experimentation with this model. Response to standard of care, sorafenib or doxorubicin, were similar to those previously reported in the literature, with however a strong toxicity of doxorubicin. Tumor vascularization was visible by histology seven days after Huh-7-Luc transplantation and robustly developed at day 14 and day 21. The model was used to explore different imaging modalities, including microtomography, probe-based confocal laser endomicroscopy, full-field optical coherence tomography, and ultrasound imaging. Tumor engraftment was similar after echo-guided intrahepatic injection as after laparotomy. Collectively, this orthotopic hepatocellular carcinoma model enables the in vivo evaluation of chemotherapeutic and surgical approaches using multimodal imaging.

Development of a methodology for in vivo follow-up of hepatocellular carcinoma in hepatocyte specific Trim24-null mice treated with myo-inositol trispyrophosphate.

BACKGROUND: Genetically induced hepatocellular carcinoma (HCC) models are generally used to investigate carcinogenesis pathways, but very few attempts were made to valorize them for pharmacological testing. This study describes a micro-computed tomography (micro-CT) - based methodology for the diagnostic and lifelong follow-up of HCC in the hepatocyte-specific Trim24-null mouse line. Myo-inositol trispyrophosphate (ITPP) was tested as anti-cancer drug. METHODS: Partial hepatectomy was performed in 2 months-old Trim24-null mice, in order to accelerate the carcinogenesis process. HCC diagnosis was obtained by micro-CT scan with double contrast agent: 10 µl/g Fenestra™ LC was injected intraperitoneally 6 h prior to imaging and 10 µl/g Fenestra™ VC was injected intravenously 15 min prior to imaging. Twenty three hepatocyte-specific Trim24-null mice were considered for ITPP testing (3 mg/g/week intraperitoneally during 10 months in 12 mice, versus 11 controls). Lifelong follow-up was performed using micro-CT. Comparative analysis was performed using unpaired t test with Welch correction and survival curves were compared by log-rank test. Gene expression analysis was performed using the RT q-PCR technique. RESULTS: Double contrast micro-CT scan allowed HCC diagnosis as hypodense, isodense or hyperdense nodules. Positive predictive value was 81.3 %. Negative predictive value was 83.3 %. Tumor growth could be objectified by micro-CT scan before the ITPP treatment was started, and at 3 and 9 months follow-up. Significant progression of tumor volume was demonstrated in the both groups, with no difference between groups (p > 0.05). In the ITPP group, a mild decrease in tumor doubling time was first observed (31.9 +/- 12 days, p > 0.05) followed by a significant increase (59.8 +/- 18.3 days, p = 0.008). However, tumor doubling time was not different between groups (p > 0.05). Median survival after treatment initiation was 223 days (controls) versus 296 days (ITPP group, p = 0.0027). HIF1α, VEGF, glutamine synthase, osteopontin expression levels were not significantly modified at the end of follow-up. In the ITPP group, the p53 expression profile was inversed as compared to the control group, higher in non-tumor livers than in tumors. CONCLUSION: ITPP treatment allowed for a two-month survival improvement, with better tolerance of tumor burden and apoptosis increase in non-tumor, pathological livers.

Transcription cofactors TRIM24, TRIM28, and TRIM33 associate to form regulatory complexes that suppress murine hepatocellular carcinoma.

TRIM24 (TIF1α), TRIM28 (TIF1ß), and TRIM33 (TIF1γ) are three related cofactors belonging to the tripartite motif superfamily that interact with distinct transcription factors. TRIM24 interacts with the liganded retinoic acid (RA) receptor to repress its transcriptional activity. Germ line inactivation of TRIM24 in mice deregulates RA-signaling in hepatocytes leading to the development of hepatocellular carcinoma (HCC). Here we show that TRIM24 can be purified as at least two macromolecular complexes comprising either TRIM33 or TRIM33 and TRIM28. Somatic hepatocyte-specific inactivation of TRIM24, TRIM28, or TRIM33 all promote HCC in a cell-autonomous manner in mice. Moreover, HCC formation upon TRIM24 inactivation is strongly potentiated by further loss of TRIM33. These results demonstrate that the TIF1-related subfamily of TRIM proteins interact both physically and functionally to modulate HCC formation in mice.

Cell death after high-LET irradiation in orthotopic human hepatocellular carcinoma in vivo.

Hepatocellular carcinoma (HCC) represents the sixth most common cancer worldwide and a major health problem since the choice of treatment is limited due to chemo- and radio-resistance. It was previously reported that high linear energy transfer (LET) radiation induced massive autophagic cell death in the human HCC SK-Hep1 cell line in vitro. This study analyzed the effects of high-LET radiation on the same HCC tumor model, orthotopically transplanted into nude mice. For this purpose, after surgical xenograft in the liver, animals were irradiated with fast neutrons and cell death occurring in the tumors was assessed with various techniques, including electron microscopy and probe-based confocal laser endomicroscopy. Results indicate that considerable autophagy and only limited apoptosis took place in the tumor xenografts after high-LET irradiation. These data confirm the previous in vitro results, suggesting that autophagy may act as a predominant mode of cell death in the efficacy of high-LET radiation.

Pharmacological enhancement of autophagy induced in a hepatocellular carcinoma cell line by high-LET radiation.

The aim of the present study was to determine the cytotoxic consequences of high-linear energy transfer (LET) irradiation in the presence of oxaliplatin on hepatocellular carcinoma (HCC) cells in vitro. We attempted to correlate the induction of apoptosis and autophagy with the formation of DNA double-strand breaks (DSBs). SK-Hep1 cells were irradiated by 65 MeV neutrons in the presence of oxaliplatin and/or the poly(ADP-ribose) polymerase (PARP) inhibitor PJ34. DSBs were measured by the formation of gammaH2AX foci. Results show that in SK-Hep1 cells exposed to fast neutrons in the presence of oxaliplatin, DSBs occurred and persisted with time after irradiation. While apoptosis remained low in co-treated cells, autophagy was considerably increased after irradiation and augmented by the addition of oxaliplatin. Thus, autophagic cell death appears to play a prominent role in the cytotoxicity of the combined treatment and may be linked to the generation of heavy damage to DNA.

Feasibility and reliability of pancreatic cancer staging using fiberoptic confocal fluorescence microscopy in rats.

BACKGROUND & AIMS: Surgical management of pancreatic cancer depends on tumor resectability and staging. This study evaluated a new in vivo technique, fiberoptic confocal fluorescence microscopy (FCFM), for detection and staging of pancreatic tumors in rats. METHODS: FCFM was used with a protease-activated fluorescent marker (ProSense; VisEn Medical Inc, Woburn, MA) for in vivo imaging of solid organs (1.8-microm resolution) in a rat model of pancreatic ductal adenocarcinoma. A preliminary study described the FCFM rendering of normal and pathologic tissues. Subsequently, 2 double-blind studies compared FCFM to standard histology in (1) detection of tumors in rat models of cancer and controls and (2) detection of nodal involvement (splenic, celiac, mesenteric, and colic) 4, 5, and 6 weeks after tumor induction vs controls. RESULTS: Tumor cells displayed a fluorescent ductal pattern compared with non-fluorescent normal pancreas or normal follicular pattern of lymph nodes (LNs). FCFM detected all the pancreatic tumors (1.7-mm mean diameter) and identified 23 LNs that contained metastases of 99 LNs examined. Standard histologic analyses resulted in 1 false-negative result in tumor detection and 2 false negatives in LN detection, whereas FCFM produced no false-negative results. Additional serial sectioning confirmed all tumors and 16 metastatic LNs; FCFM had a negative predictive value of 100% and a positive predictive value of 69.6%. CONCLUSIONS: Real-time "virtual biopsy" using FCFM detects tumors and LN metastases with 100% sensitivity and 92.2% specificity in rats, making it a reliable technique for detection and staging of pancreatic cancer.

Arterial calcifications and increased expression of vitamin D receptor targets in mice lacking TIF1alpha.

Calcification of arteries is a major risk factor for cardiovascular mortality in humans. Using genetic approaches, we demonstrate here that the transcriptional intermediary factor 1alpha (TIF1alpha), recently shown to function as a tumor suppressor in murine hepatocytes, also participates in a molecular cascade that prevents calcifications in arterioles and medium-sized arteries. We further provide genetic evidence that this function of TIF1alpha is not exerted in hepatocytes. The sites of ectopic calcifications in mutant mice lacking TIF1alpha resemble those seen in mice carrying an activating mutation of the calcium sensor receptor (Casr) gene and, in TIF1alpha-deficient kidneys, Casr expression is increased together with that of many other vitamin D receptor (VDR) direct target genes, namely Car2, Cyp24a1, Trpv5, Trpv6, Calb1, S100g, Pthlh, and Spp1. Thus, our data indicate that TIF1alpha represses the VDR pathway in kidney and suggest that an up-regulation of Casr expression in this organ could account for ectopic calcifications generated upon TIF1alpha deficiency. Interestingly, the calcifying arteriopathy of TIF1alpha-null mutant mice shares features with the human age-related Mönckeberg's disease and, overall, the TIF1alpha-null mutant pathological phenotype supports the hypothesis that aging is promoted by increased activity of the vitamin D signaling pathway.

Loss of Trim24 (Tif1alpha) gene function confers oncogenic activity to retinoic acid receptor alpha.

Hepatocellular carcinoma (HCC) is a major cause of death worldwide. Here, we provide evidence that the ligand-dependent nuclear receptor co-regulator Trim24 (also known as Tif1alpha) functions in mice as a liver-specific tumor suppressor. In Trim24-null mice, hepatocytes fail to execute proper cell cycle withdrawal during the neonatal-to-adult transition and continue to cycle in adult livers, becoming prone to a continuum of cellular alterations that progress toward metastatic HCC. Using pharmacological approaches, we show that inhibition of retinoic acid signaling markedly reduces hepatocyte proliferation in Trim24-/- mice. We further show that deletion of a single retinoic acid receptor alpha (Rara) allele in a Trim24-null background suppresses HCC development and restores wild-type expression of retinoic acid-responsive genes in the liver, thus demonstrating that in this genetic background Rara expresses an oncogenic activity correlating with a dysregulation of the retinoic acid signaling pathway. Our results not only provide genetic evidence that Trim24 and Rara co-regulate hepatocarcinogenesis in an antagonistic manner but also suggest that aberrant activation of Rara is deleterious to liver homeostasis.