Alexander Shtabsky

THE EFFECT OF PNEUMOPERITONEUM ON DISSEMINATION AND SCAR IMPLANTATION OF INTRA-ABDOMINAL TUMOR CELLS

PURPOSE The role of laparoscopy for the treatment of cancer remains controversial, and a particular concern is port site metastases after laparoscopic surgery. Since laparoscopy is being performed with increasing frequency, the question arises as to whether it is a safe oncological procedure. After intraperitoneal inoculation of renal cell carcinoma cells in a mouse model, we compare abdominal wall scar implantation following laparoscopic trocar insertion and pneumoperitoneum with standard laparotomy, and examine the effects on tumor dissemination in the peritoneal cavity. MATERIALS AND METHODS Following intra-abdominal RENCA cell inoculation, Balb/c mice were randomized into group 1-20 mice that underwent carbon dioxide pneumoperitoneum and telescope trocar insertion, group 2-20 subjected to laparotomy and group 3-10 anesthetized only. All animals were sacrificed 2 weeks after inoculation, and abdominal wall metastases and intraperitoneal tumor distribution were evaluated. RESULTS Overall, intra-abdominal implantation of inoculated RENCA tumor cells was detected in 15 of 20 animals (75%) in group 1, 14 of 20 (70%) in group 2 and 10 of 10 (100%) in group 3. Wound metastases developed in 46.7% of the mice in group 1 and 50% in group 2. CONCLUSIONS There was no difference among the groups in the pattern of intraperitoneal tumor implants and scar seeding incidence. Pneumoperitoneum does not facilitate port site metastases.

Embryonic Porcine Liver as a Source for Transplantation: Advantage of Intact Liver Implants over Isolated Hepatoblasts in Overcoming Homeostatic Inhibition by the Quiescent Host Liver

Cell therapy as an alternative to orthotopic liver transplantation represents a major challenge, since negligible proliferation of isolated hepatocytes occurs after transplantation because of the stringent homeostatic control displayed by the host liver. Thus, different modalities of liver injury as part of the pretransplant conditioning are a prerequisite for this approach. The major objective of the present study was to test whether xenotransplantation of pig fetal liver fragments, in which potential cell‐cell and cell‐stroma interactions are spared, might afford more robust growth and proliferation compared with isolated pig fetal hepatoblasts. After transplantation into SCID mice, fetal liver tissue fragments exhibited marked growth and proliferation, in the setting of a quiescent host liver, compared with isolated fetal hepatoblasts harvested at the same gestational age (embryonic day 28). The proliferative advantage of fetal pig liver fragments was clearly demonstrated by immunohistochemical and morphometric assays and was observed not only after implantation into the liver but also into extrahepatic sites, such as the spleen and the subrenal capsule. The presence of all types of nonparenchymal liver cells that is crucial for normal liver development and regeneration was demonstrated in the implants. Preservation of the three‐dimensional structure in pig fetal liver fragments enables autonomous proliferation of transplanted hepatic cells in the setting of a quiescent host liver, without any requirement for liver injury in the pretransplant conditioning. The marked proliferation and functional maturation exhibited by the pig fetal liver fragments suggests that it could afford a preferable source for transplantation.

Rosiglitazone improves aortic arginine transport, through inhibition of PKCα, in uremic rats

Peroxisome proliferator-activated receptor (PPAR) agonists were shown to inhibit atherosclerosis through augmentation of endothelial nitric oxide synthase (eNOS) activity. In addition, rosiglitazone exerts a beneficial effect in chronic renal failure (CRF). Since l-arginine transport by CAT-1 (the specific arginine transporter for eNOS) is inhibited in uremia, we aimed to explore the effect of rosiglitazone on arginine transport in CRF. Arginine uptake by aortic rings was studied in control animals, rats, 6 wk following 5/6 nephrectomy (CRF) and rats with CRF treated with rosiglitazone. The decrease of arginine transport in CRF was prevented by rosiglitazone. Immunobloting revealed that CAT-1 protein was decreased in CRF but remained unchanged following rosiglitazone administration. Protein content of the membrane fraction of PKCalpha and phosphorylated CAT-1 increased significantly in CRF, effects that were prevented by rosiglitazone. PKCalpha phosphorylation was unchanged but significantly attenuated by rosiglitazone in CRF. Ex vivo administration of phorbol-12-myristate-13-acetate to rosiglitazone-treated CRF rats significantly attenuated the effect of rosiglitazone on arginine uptake. The decrease in cGMP response to carbamyl-choline (eNOS agonist) was significantly attenuated by rosiglitazone in CRF. Western blotting and immunohistochemistry analysis revealed that protein nitration was intensified in the endothelium of CRF rats and this was attenuated by rosiglitazone. In conclusion, rosiglitazone prevents the decrease in arginine uptake in CRF through both depletion and inactivation of PKCalpha. These findings are associated with restoration of eNO generation and attenuation of protein nitration and therefore may serve as a novel mechanism to explain the beneficial effects of rosiglitazone on endothelial function in uremia.

Targeting ErbB-1 and ErbB-4 in irradiated head and neck cancer: Results of in vitro and in vivo studies

ErbB oncogenes have a major role in cancer. The role of ErbB‐4 in cancer cell biology and the effect of anti‐ErbB‐1 and anti‐ErbB‐4 monoclonal antibodies were evaluated in this study.

Laser-induced hyperthermia for treatment of granulation tissue growth in rats

Objective: We aimed to develop a new technique for treatment of granulation tissue (GT) growth using local hyperthermia. Methods: A temperature-controlled diode laser system was developed for induction of mild hyperthermia in real time. GT was generated by harvesting the skin over the gluteal fascia in rats. Histopathological analysis was used to estimate the effect of hyperthermia on the tissue. Results: In untreated rats, GT was detected within 3 days and reached maximal thickness after 12 days. Hyperthermia at 43°C and above significantly decreased GT thickness (n = 8 per group). Hyperthermia at 48°C for 3 minutes was the most efficient parameter for treatment of GT (51% reduction), with minimal (5%) muscle necrosis. Conclusions: Hyperthermia can significantly inhibit GT growth, with minimal damage to surrounding structures. Our findings suggest a possible role for hyperthermia as a therapeutic model against GT. Further research and long-term studies are needed to explore the utility of laser-induced hyperthermia for inhibition of GT growth.

l-Arginine improves endothelial function, independently of arginine uptake, in aortas from chronic renal failure female rats

Endothelial cell dysfunction (ECD) is a common feature of chronic renal failure (CRF). Defective nitric oxide (NO) generation due to decreased endothelial nitric oxide synthase (eNOS) activity is a crucial parameter characterizing ECD. Decreased activity of cationic amino acid transporter-1 (CAT-1), the selective arginine transporter of eNOS, has been shown to inhibit eNOS in uremia. Recently, we failed to demonstrate a decrease in glomerular arginine transport in uremic female rats (Schwartz IF, Grupper A, Soetendorp H, Hillel O, Laron I, Chernichovski T, Ingbir M, Shtabski A, Weinstein T, Chernin G, Shashar M, Hershkoviz R, Schwartz D. Am J Physiol Renal Physiol 303: F396-F404, 2012). The current experiments were designed to determine whether sexual dimorphism which characterizes glomerular arginine transport system in uremia involves the systemic vasculature as well and to assess the effect of L-arginine in such conditions. Contractile and vasodilatory responses, ultrastructural changes, and measures of the L-arginine-NO system were performed in thoracic aortas of female rats subjected to 5/6 nephrectomy. The contractile response to KCl was significantly reduced, and acetylcholine-induced vasodilation was significantly impaired in aortas from CRF dames compared with healthy rats. Both of these findings were prevented by the administration of arginine in the drinking water. The decrease in both cGMP generation, a measure of eNOS activity, and aortic eNOS and phosphorylated eNOS abundance observed in CRF rats was completely abolished by l-arginine, while arginine transport and CAT-1 protein were unchanged in all experimental groups. Arginine decreased both serum levels of advanced glycation end products and the asymmetrical dimethylarginine/arginine ratio and restored the endothelial ultrastructure in CRF rats. In conclusion. arginine administration has a profound beneficial effect on ECD, independently of cellular arginine uptake, in CRF female rats.

A Novel Urine-Based Assay for Bladder Cancer Diagnosis: Multi-Institutional Validation Study

BACKGROUND CellDetect is a unique histochemical stain enabling color and morphological discrimination between malignant and benign cells based on differences in metabolic signature. OBJECTIVE The objective of the present study was to validate the performance of this assay in a controlled, blinded, multicenter study. DESIGN, SETTING, AND PARTICIPANTS The study, conducted in nine hospitals, included patients with documented history of bladder cancer, monitored for urothelial carcinoma (UCC) or scheduled for bladder cancer surgery. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS Cystoscopy and/or biopsy were used as a reference standard to determine sensitivity and specificity. Smears were stained by CellDetect and interpreted by two cytologists blinded to the patients final diagnosis. The findings were compared with those of standard urine cytology and BTA stat. RESULTS AND LIMITATIONS Two hundred and seventeen voided urine specimens were included. Ninety-six (44%) were positive by histology and 121 (56%) were negative by either cystoscopy or histology. The overall sensitivity of CellDetect was 84%. Notably, the sensitivity for detecting low-grade nonmuscle-invasive bladder cancer tumors was greater than this of BTA stat (78% vs 54%) and more than two-fold higher compared with standard cytology (33%, p ≤ 0.05). The specificity was 84% in patients undergoing routine surveillance by cystoscopy. At a median follow-up of 9 mo, 21% of the patients with positive CellDetect and negative reference standard developed UCC, which was significantly higher compared with the 5% of the true negative cases. Limitations include the lack of instrumental urine samples and the lack of patients with nongenitourinary cancers in the study population. CONCLUSIONS This study validates the performance of CellDetect as a urine-based assay to identify UCC in patients with history of bladder cancer. The high sensitivity was maintained across all cancer grades and stages without compromising the assay specificity. Further studies are required to test whether this novel stain can be incorporated in routine bladder cancer surveillance as a noninvasive alternative to cystoscopy. PATIENT SUMMARY Surveillance of bladder cancer requires frequent invasive procedures. In the present study, we validate the ability of a novel biomarker to accurately identify early-stage tumors in urine specimens for the noninvasive monitoring of patients with history of bladder cancer.

Angioimmunoblastic T-cell lymphoma presenting as giant kidneys: a case report

IntroductionAngioimmunoblastic T-cell lymphoma is a rare form of tumor of the lymph nodes or lymphoid tissue. In this report we describe an unusual presentation of angioimmunoblastic T-cell lymphoma consisting of giant kidneys with no nephrotic syndrome.Case presentationA 46-year-old Arabic man from Gaza was hospitalized in our ward due to abdominal pain and a weight loss of 20 kg during the preceding two months. The results of the physical examination and laboratory tests raised the possibility of neoplastic disease. A computerized tomographic scan of the abdomen showed huge kidneys, and a kidney biopsy showed infiltration by lymphocytes and eosinophils. The genetic examination revealed T-cell lymphoma. Diagnosis was made by a lymph node biopsy, which shows typical findings of angioimmunoblastic T-cell lymphoma.ConclusionsAngioimmunoblastic T-cell lymphoma can present with huge kidneys without nephrotic syndrome.

Combined acetyl-11-keto-β-boswellic acid and radiation treatment inhibited glioblastoma tumor cells

Glioblastoma multiforme (GBM) is the most common and most aggressive subtype of malignant gliomas. The current standard of care for newly diagnosed GBM patients involves maximal surgical debulking, followed by radiation therapy and temozolomide chemotherapy. Despite the advances in GBM therapy, its outcome remains poor with a median survival of less than two years. This poor outcome is partly due to the ability of GBM tumors to acquire adaptive resistance to therapy and in particular to radiation. One of the mechanisms contributing to GBM tumor progression and resistance is an aberrant activation of NF-ĸB, a family of inducible transcription factors that play a pivotal role in regulation of many immune, inflammatory and carcinogenic responses. Acetyl-11-keto-β-boswellic acid (AKBA) is a pentacyclic terpenoid extracted from the gum Ayurvedic therapeutic plant Boswellia serrata. AKBA is anti-inflammatory agent that exhibits potent cytotoxic activities against various types of tumors including GBM. One of the mechanisms underlying AKBA anti-tumor activity is its ability to modulate the NF-ĸB signaling pathway. The present study investigated in vitro and in vivo the effect of combining AKBA with ionizing radiation in the treatment of GBM and assessed AKBA anti-tumor activity and radio-enhancing potential. The effect of AKBA and/or radiation on the survival of cultured glioblastoma cancer cells was evaluated by XTT assay. The mode of interaction of treatments tested was calculated using CalcuSyn software. Inducing of apoptosis following AKBA treatment was evaluated using flow cytometry. The effect of combined treatment on the expression of PARP protein was analysed by Western blot assay. Ectopic (subcutaneous) GBM model in nude mice was used for the evaluation of the effect of combined treatment on tumor growth. Immunohistochemical analysis of formalin-fixed paraffin-embedded tumor sections was used to assess treatment-related changes in Ki-67, CD31, p53, Bcl-2 and NF-ĸB-inhibitor IĸB-α. AKBA treatment was found to inhibit the survival of all four tested cell lines in a dose dependent manner. The combined treatment resulted in a more significant inhibitory effect compared to the effect of treatment with radiation alone. A synergistic effect was detected in some of the tested cell lines. Flow cytometric analysis with Annexin V-FITC/PI double staining of AKBA treated cells indicated induction of apoptosis. AKBA apoptotic activity was also confirmed by PARP cleavage detected by Western blot analysis. The combined treatment suppressed tumor growth in vivo compared to no treatment and each treatment alone. Immunohistochemical analysis showed anti-angiogenic and anti-proliferative activity of AKBA in vivo. It also demonstrated a decrease in p53 nuclear staining and in Bcl-2 staining and an increase in IĸB-α staining following AKBA treatment both alone and in combination with radiotherapy. In this study, we demonstrated that AKBA exerts potent anti-proliferative and apoptotic activity, and significantly inhibits both the survival of glioblastoma cells in vitro and the growth of tumors generated by these cells. Combination of AKBA with radiotherapy was found to inhibit factors which involved in cell death regulation, tumor progression and radioresistence, therefore it may serve as a novel approach for GBM patients.

A novel assessment of the quality of immunohistostaining overcomes the limitations of current methods.

Quality assurance has become an integral part of surgical pathology. Despite the development of interdisciplinary quality systems, however, the means for objective analysis in surgical pathology are limited. Immunohistostaining is a multi-factorial procedure that depends on the quality of reagents and antibodies employed in the process and on technical methodology. In the present study, we aim to establish a straightforward procedure for objective quality evaluation of the components involved in immunohistostaining. The quality of two of these components, the primary antibody and the automated staining device, was assessed by employing each component from two different sources, one serving as the test substance and the second as the reference. Assessment was performed by at least two pathologists in a blinded fashion using pre-established quality criteria and scores. The quality analysis of two automated devices revealed a significant difference between the reference and tested devices (3.5+/-1.7 and 4.2+/-1.5, respectively, P>0.05), while the analysis of two selected antibodies did not reveal any statistical difference. The described method provided objective quality assessment of selected components affecting immunohistostaining by elaborating numeric values that enabled statistical analysis. This approach is applicable to any given component in various surgical pathology procedures.