Characterization of Transplantable Insulinoma Cells.

This chapter describes the propagation and characterization of transplantable insulinoma cells as model of insulin-producing pancreatic islet cells in the rat. Here, the cells are propagated by transplantation into rats followed by harvesting after growth for approximately 1 month. The cells are then purified by Percoll density gradient centrifugation and characterized by pulse-chase radiolabelling and immunoprecipitation of the insulin-related peptides. The results show that the transplantable insulinoma cells produce insulin in a manner similar to that found in normal pancreatic islets.

Hipoglucemia post cirugía bariátrica. Presentación de un caso con buena respuesta a octreótide / HYPOGLYCEMIA COMPLICATING BARIATRIC SURGERY. CASE REPORT WITH SUCCESSFUL RESPONSE TO OCTROTIDE

El Bypass Gástrico (BPG) representa la herramienta terapéutica más efectiva para el manejo de la obesidad. Sin embargo, la hipoglucemia con neuroglucopenia post BPG es una complicación que se describe cada vez con mayor frecuencia. Se presenta el caso de una paciente con hipoglucemia hiperinsulinémica (HH) post BPG y los distintos esquemas terapéuticos utilizados, pudiendo controlar finalmente los valores de glucemia con octreótide y evitando así, la realización de una pancreatectomía para el tratamiento de las hipoglucemias

Gastric Bypass (GBP) is the most effective treatment for patients with severe obesity. Hyperinsulinemic hypoglycemia with neuroglycopenia is an increasingly late complication of GBP. A case of a post GBP hyperinsulinemic hypoglycemia is reported, and the different drugs used for its treatment, being able to control the glycemia with octreotide and avoiding a pancreatectomy surgery as hypoglycemia treatment

A freeze-dried kit formulation for the preparation of Lys(27)(99mTc-EDDA/HYNIC)-Exendin(9-39)/99mTc-EDDA/HYNIC-Tyr3-Octreotide to detect benign and malignant insulinomas.

About 90% of insulinomas are benign and 5%-15% are malignant. Benign insulinomas express the glucagon-like peptide-1 receptor (GLP-1R) and low levels of somatostatin receptors (SSTR), while malignant insulinomas over-express SSTR or GLP-1R in low levels. A kit for the preparation of Lys(27)((99m)Tc-EDDA/HYNIC)-Exendin(9-39)/(99m)Tc-EDDA/HYNIC-Tyr(3)Octreotide was formulated to detect 100% of insulinomas. The formulation showed radiochemical purity of 97±1%, high stability in human serum, and GLP-1R and SSTR affinity. The biodistribution and imaging studies demonstrated properties suitable for its use as a target-specific agent for the simultaneous molecular imaging of GRP-1R- and/or SSTR-positive tumors.

Transformation of nonfunctioning pancreatic tumor into malignant insulinoma after 3 years: an uncommon clinical course of insulinoma.

A 62-year-old man admitted to our outpatient clinic with two months of recurrent life threatening hypoglycemia episodes. He was diagnosed as malignant insulinoma with multiple metastases of liver and peripancreatic lymph nodes. Liver biopsy specimen was demonstrated grade 2 neuroendocrine tumor compatible with clinical and radiological results. He was followed under the treatment of continuous intravenous glucose infusion during the diagnostic procedures. He had a pancreatic lesion history measured 20 x 12 mm in diameter via the abdominal tomography examination approximately two years before the diagnosis. Unusual course of this case suggests the transformation of nonfunctioning pancreatic neuroendocrine tumor into functional insulin secreting tumor with metastases. The patient was found inoperable and started on chemotherapy.

Giant insulinoma: a report of 3 cases and review of the literature.

Insulinoma is a rare pancreatic neuroendocrine tumor that is usually described as benign, sporadic, and very small (<2 cm). However, there have been rare case reports of insulinoma presenting as a giant tumor. We describe 3 cases of giant insulinomas, all of which developed liver metastases. The patients were aged 38, 63, and 67 years. Clinically, all patients presented with Whipple's triad associated with a large mass located in the pancreatic tail. The tumors ranged in size from 10 to 15 cm. On microscopic examination, the tumors were well differentiated with amyloid deposition ranging between 20% and 30%. Immunohistochemically, all 3 tumors showed strong diffuse expression of chromogranin and synaptophysin, whereas they were only focally positive for insulin. One patient developed liver recurrence 3 years after resection of the primary tumor yet remained asymptomatic without treatment. Another patient with liver recurrence underwent right hepatectomy and has been free of disease for 2 years. The third patient died of metastatic disease 13 years after initial surgery. Giant insulinomas are characterized by focal expression of insulin and high rates of liver metastases. Long-term follow-up is mandatory in these patients, as recurrence is expected after primary surgery.

Potential contribution of translational factors to triiodo-L-thyronine-induced insulin synthesis by pancreatic beta cells.

BACKGROUND: Thyroid hormones (THs) are known to regulate protein synthesis by acting at the transcriptional level and inducing the expression of many genes. However, little is known about their role in protein expression at the post-transcriptional level, even though studies have shown enhancement of protein synthesis associated with mTOR/p70S6K activation after triiodo-L-thyronine (T3) administration. On the other hand, the effects of TH on translation initiation and polypeptidic chain elongation factors, being essential for activating protein synthesis, have been poorly explored. Therefore, considering that preliminary studies from our laboratory have demonstrated an increase in insulin content in INS-1E cells in response to T3 treatment, the aim of the present study was to investigate if proteins of translational nature might be involved in this effect. METHODS: INS-1E cells were maintained in the presence or absence of T3 (10(-6) or 10(-8) M) for 12 hours. Thereafter, insulin concentration in the culture medium was determined by radioimmunoassay, and the cells were processed for Western blot detection of insulin, eukaryotic initiation factor 2 (eIF2), p-eIF2, eIF5A, EF1A, eIF4E binding protein (4E-BP), p-4E-BP, p70S6K, and p-p70S6K. RESULTS: It was found that, in parallel with increased insulin generation, T3 induced p70S6K phosphorylation and the expression of the translational factors eIF2, eIF5A, and eukaryotic elongation factor 1 alpha (eEF1A). In contrast, total and phosphorylated 4E-BP, as well as total p70S6K and p-eIF2 content, remained unchanged after T3 treatment. CONCLUSIONS: Considering that (i) p70S6K induces S6 phosphorylation of the 40S ribosomal subunit, an essential condition for protein synthesis; (ii) eIF2 is essential for the initiation of messenger RNA translation process; and (iii) eIF5A and eEF1A play a central role in the elongation of the polypeptidic chain during the transcripts decoding, the data presented here lead us to suppose that a part of T3-induced insulin expression in INS-1E cells depends on the protein synthesis activation at the post-transcriptional level, as these proteins of the translational machinery were shown to be regulated by T3.

Insulinoma diagnosed in the postpartum: clinical and immunohistochemical features.

Insulinomas are rare pancreatic ß-cell tumors with an estimated incidence of 1:250.000 persons/year. We present a novel case of insulinoma manifesting immediately after childbirth. Eight days after delivery, a 21-year-old, previously healthy woman presented paresthesia in hands, upper and lower limbs muscle weakness with difficult walking, which worsened during breastfeeding sessions. Laboratory tests showed blood glucose levels between 37 and 55 mg/dL with inappropriately normal insulin levels (7.78 µUI/mL; normal range: 5-29). An abdominal computed tomography showed a nodular lesion measuring 2 cm at the head of the pancreas. Tumor enucleation resulted in complete resolution of hypoglycemia. Histopathological and immunohistochemical analysis were consistent with an insulinoma. About 27 cases of insulinoma associated with pregnancy have been reported to date, mostly diagnosed before the 16th week. The beginning of symptoms soon after delivery is less common. Understanding the interactions between pancreatic ß-cell function and all the physiological metabolic and hormonal adaptations associated with gestation is essential for the adequate management of hypoglycemic disorders in pregnant women.

Pancreatic insulinoma: a surgical experience.

BACKGROUND: Small size, high benignity rate, and sporadic nature make insulinomas suitable for laparoscopic resection. On the other hand, occult location or multicentricity mandate open surgery. This study was designed to analyze a series of patients who had pancreatic insulinomas and underwent initial treatment at our institution. METHODS: Clinical records of the 34 patients with pancreatic insulinomas who underwent surgical resection between 1995 and 2007 were reviewed. Main variables for analysis were cure of the disease and surgical complications. RESULTS: There were 20 women and 14 men with a mean age of 40 +/- 13 years. Mean size of the tumors was 2.2 +/- 1 cm. Laparoscopic resection was completed in 14 of 21 patients. Most tumors that were resected by laparoscopy were solitary, benign, and located in the body and tail of the pancreas. Open surgery was selected for 13 patients, including 7 sporadic (5 in the head), 4 related to the MEN syndrome, and 2 malignant tumors. Surgical morbidity occurred in 23 patients. The most common complication was pancreatic fistula (3/13 in open, 4/14 in laparoscopic, and 6/7 in conversions). One patient in the open group died 15 days after surgery from massive PTE. Postoperative normoglycemia was achieved in all patients and persisted for a follow-up period of 4 +/- 3.7 years. CONCLUSIONS: Most insulinomas in our series were small and benign. Tumors that were located in the body and tail were more often amenable for laparoscopic resection. The cure rate was very high. Pancreatic fistula was the most frequent complication.

Malignant insulinoma arising from intrasplenic heterotopic pancreas.

CONTEXT: Heterotopic pancreas is defined as ectopic pancreatic tissue without vascular or anatomic continuity with the normal pancreas. The spleen is a rare site of origin. This case report describes a patient with a malignant insulinoma which originated from an intrasplenic heterotopic pancreas. CASE REPORT: A 46-year-old man with three previous episodes of neuroglucopenic and adrenergic symptoms was referred to our hospital. A fasting test was performed and discontinued due to hypoglycemic symptoms. Preoperative studies failed to demonstrate any pancreatic lesions. However, a heterogeneous encapsulated tumor in the spleen was found on MRI. During surgery, only the splenic tumor was found, with neither vascular nor anatomical connections to the normal pancreas. Pathology reported a malignant insulinoma. Insulin and proinsulin were documented by immunohistochemistry. After one year of follow up, the patient is free of symptoms and no recurrent disease has been documented. DISCUSSION: Only seven cases of splenic heterotopic pancreas have been reported, six with cystic mucinous neoplasms. In addition, only one case of a malignant insulinoma arising from heterotopic pancreas has previously been described. This is the second case reported of an insulinoma arising from heterotopic pancreas and the first to originate from intrasplenic heterotopia.

Pancreatic neuroendocrine tumours.

INTRODUCTION: Pancreatic neuroendocrine tumours (PNT) are infrequent epithelial neoplasms associated with a better outcome than pancreatic adenocarcinoma. MATERIALS AND METHODS: We analysed our 22 years of experience in managing PNT. Forty-nine patients (27 women and 22 men) with a mean age of 49 years were studied. There were 28 insulinomas, eight glucagonomas, three gastrinomas, one VIPoma and one carcinoid. Eight patients presented with nonfunctional tumours. Enucleation was performed in 20 patients, distal pancreatectomy in 16, middle pancreatic resection in four, cephalic pancreatoduodenectomy in two and total pancreatoduodenectomy in one. In six patients, the tumour was not resected. RESULTS: Postoperative complication rate was 22%: six pancreatic fistulas, three intra-abdominal collections, one remnant pancreatitis and one pancreatic pseudocyst. There was no mortality. 39 cases showed benign histologic features and ten malignant ones. Symptomatic palliation was achieved in 94% of the cases. Five patients presented recurrences: three liver metastases and two pancreatic recurrences. Actuarial mean survival was 163 months and was longer in insulinomas, in those tumours completely resected and in tumours with benign histological features. CONCLUSION: Conservative surgery of the pancreas is preferred, but aggressive surgery is indicated when the primary tumour can be controlled. Despite of minimising pancreatic resection, there is a high complication rate, mainly pancreatic fistulas, though they can often be conservatively managed. Insulinomas are the PNT with better outcome; those completely resected also associate a better prognosis.

Serpin peptidase inhibitor clade A member 1 as a potential marker for malignancy in insulinomas.

PURPOSE: The biological behavior of insulinomas cannot be predicted based on histopathologic criteria in which the diagnosis of malignancy is confirmed by the presence of metastases. In this study, microarray and quantitative real-time reverse transcription-PCR were applied to identify differentially expressed genes between malignant and nonmalignant insulinomas to search for useful biomarkers to recognize the metastatic potential of insulinomas. EXPERIMENTAL DESIGN: Code Link human bioarrays were used to analyze differences in approximately 20,000 genes between six well-differentiated endocrine tumors of benign behavior compared with one well-differentiated endocrine carcinoma (WDEC) and three metastases of endocrine carcinomas (MEC). Quantitative real-time reverse transcription-PCR was used to validate differential expressions of five genes in a series of 35 sporadic insulinomas. Serpin peptidase inhibitor clade A member 1 (SERPINA1; alpha-1-antitrypsin) expression, identified as up-regulated in malignant insulinomas, was also evaluated by immunohistochemistry. RESULTS: Analysis of microarray data resulted in 230 differentially expressed genes. Gene Ontology analysis identified serine-type endopeptidase activity and serine-type endopeptidase inhibitor activity as pathways presenting significant differential expression. Protease serine 2 and complement factor B (from serine-type endopeptidase activity pathway) were respectively confirmed as up-regulated in well-differentiated endocrine tumors of benign behavior (WDET) and in WDEC/MEC. Angiotensinogen and SERPINA1 (from serine-type endopeptidase inhibitor activity pathway) were confirmed as up-regulated in WDEC/MEC. SERPINA1 was shown to be expressed in 85.7% of malignant versus 14.3% of nonmalignant insulinomas by immunohistochemistry. CONCLUSIONS: Our data are consistent to the possibility that SERPINA1 is a marker of malignancy in insulinomas. Given the widespread availability of antibody anti-alpha-1-antitrypsin in pathology services, SERPINA1 expression evaluation might be of clinical utility in recognizing patients more likely to develop an aggressive presentation.

Effect of iNOS and NF-kappaB gene silencing on beta-cell survival and function.

PURPOSE: Type I diabetes results from beta-cell death and dysfunction, induced by infiltration of immune cells and local production of inflammatory cytokines. Therefore, we investigated the effect of iNOS and NF-kappaB gene silencing on beta-cell survival and function. METHODS: Rat insulinoma INS-1E cells were transfected with chemically synthesized siRNA after complex formation with Lipofectamine 2000. Cells were then treated with a cocktail of inflammatory cytokines (IL-1beta+ TNF-alpha+ IFN-alpha), and glucose stimulated-insulin response and viability were determined. iNOS and NF-kappaB gene expression was assessed at mRNA level by real time RT-PCR. The effect of gene silencing was also correlated with cytokine-induced NO production and apoptosis. RESULTS: Transfection of INS-1E cells with siRNAs silenced iNOS and NF-kappaB gene expression and reduced NO production in a sequence-specific manner without causing significant loss of cell viability and function. However, the abrogation of NO production did not prevent INS-1E cells from cytokine-induced apoptosis, suggesting that this event may not be totally dependent on NO production. CONCLUSION: The gene silencing approach presented here is capable of attenuating the effects of inflammatory cytokines, such as iNOS expression and NO production and it will help to identify new target genes to improve islet transplantation.

Avaliação dos insulinomas pela ultra-sonografia intra-operatória: estado atual do tema / Intraoperative ultrasonographic evaluation of insulinomas: an update

Os autores fazem uma revisão da literatura sobre a utilização da ultra-sonografia intra-operatória para a avaliação dos insulinomas pancreáticos. São referidos os resultados da ultra-sonografia intra-operatória, ultra-sonografia e tomografia computadorizada realizadas no pré-operatório, e os resultados da inspeção e palpação do pâncreas realizadas durante procedimentos cirúrgicos referidos na literatura.

The authors review the literature about intraoperative ultrasonography for evaluation of pancreatic insulinomas. Results of intraoperative ultrasound, preoperative ultrasound and computed tomography are discussed, as well as results of inspection and palpation of the pancreas during surgery, reported in the literature.

[Pancreatic insulinomas]. / Insulinoma de páncreas.

Insulinoma is the most frequent neuroendocrine pancreatic tumor. In the present study, the clinical and immunohistochemical results of 20 patients who underwent surgery between January 1986 and December 2004 were evaluated. Clinical presentation, laboratory data, imaging studies, aspects of the surgical technique, complication rates and medium- and long-term follow-up were analyzed. Surgical treatment was recommended in all patients based on presenting symptoms and laboratory signs of hypoglycemia and hyperinsulinemia. In 15 patients, the lesion was identified preoperatively. In the 5 remaining patients, intraoperative palpation and ultrasonography were used to locate the lesion. The most frequently performed surgical procedures were pancreatic resection in 10 patients and laparotomic enucleation in the remaining 10. Laparoscopy was used in two patients. Two patients developed diabetes mellitus. The most frequent surgical complication was pancreatic fistula. No mortality was observed in the present series. Symptom reversion, characterized by disappearance of Whipple's triad and normal or increased glycemia values compared with preoperative values, was observed in all patients.

Somatostatin receptor subtype 5 (SSTR5) mRNA expression is related to histopathological features of cell proliferation in insulinomas.

Insulinomas are rare endocrine neoplasias that constitute the most frequent islet cell tumours. Somatostatin (SST) analogs are tentatively used to inhibit insulin secretion and control tumour growth in patients with local invasion or inoperative metastasis, but variable responses have been reported. Data regarding somatostatin receptor (SSTR) subtypes expression in insulinomas are conflicting. In this study, we evaluated 16 cases of primary insulinomas (including four primary plurihormonal tumours) and two hepatic metastases. Histopathological and immunohistochemical analysis for some features associated with tumour aggressiveness and semi-quantitative RT-PCR for SSTR1-5 and real-time qPCR for SSTR5 were performed. SSTR subtypes 1, 3, and 5 were expressed in 100%, SSTR2 in 89%, and SSTR4 only in 22% of the insulinomas. SSTR5 mRNA was positively correlated with histopathological features related to tumour aggressiveness (large tumour diameter, well-differentiated endocrine tumour with uncertain behaviour and higher number of cells with nuclear atypia). SSTR5 mRNA expression in primary insulinomas was lower than in primary plurihormonal tumours (P < 0.05). The observed positive correlation between SSTR5 expression and tumour size suggests that the use of SST analogues more specific to SSTR5 in the treatment of insulinomas deserves attention.

Fatty acid-induced toxicity and neutral lipid accumulation in insulin-producing RINm5F cells.

Fatty acids have been shown to cause death of rat and human primary pancreatic beta cells and of insulin-producing cell lines. These studies focused mainly on saturated and monounsaturated FA such as palmitic, stearic and oleic acids. In this study, we have performed a comparison of the toxicity of a wider range of FA. The toxicity of different FA to insulin-producing RINm5F cells was assessed by flow cytometry measuring loss of plasma membrane integrity and increase in DNA fragmentation. Additionally, the FA induced neutral lipid accumulation and the FA composition were determined. Palmitic, linoleic, gamma-linolenic, oleic, stearic, and eicosapentaenoic acid caused DNA fragmentation of insulin-producing RINm5F cells. Loss of membrane integrity was mainly caused by linoleic and gamma-linolenic acid. There was no correlation between cytotoxicity and the abundance of the FA in the cells as determined by HPLC analysis. Taken as whole, the toxic effect of the FA on insulin-producing RINm5F cells varied irrespective of the chain length and the degree of unsaturation. In these cells PA and LA exhibited the highest toxicity, whereas AA was not toxic. In addition, the toxicity of most tested FA was inversely related to low NLA, except for AA and EPA. The results of this study contribute to the understanding of the role of FA in the impairment of pancreatic beta cell function that occurs in type 2 diabetes and obesity.

Cellular proliferative fraction measured with topoisomerase IIalpha predicts malignancy in endocrine pancreatic tumors.

CONTEXT: Endocrine pancreatic tumors (EPTs) are rare lesions with varying biological behavior. Establishing malignancy is a challenge for clinicians and pathologists. OBJECTIVE: To establish the role of proliferative, apoptotic, angiogenic, and hormonal markers as predictors of malignancy in EPTs. DESIGN: Paraffin-embedded EPT samples were studied for prognostic markers. PATIENTS: Twenty-one consecutive patients with a diagnosis of EPT. MAIN OUTCOME MEASURES: The proliferative fraction (topoisomerase IIalpha), microvascular density (CD34), vascular endothelial growth factor expression, and estrogen receptor-beta (ERbeta) expression were studied by immunohistochemistry on all EPTs. Apoptosis was also assessed with terminal deoxynucleotidyl transferase nick-end labeling. RESULTS: We identified 13 benign and 8 malignant tumors. Topoisomerase IIalpha was significantly increased in malignant tumors (P =.001), while there were no differences in apoptosis, microvascular density, or vascular endothelial growth factor expression in association with malignancy. No correlation could be identified between microvascular density and vascular endothelial growth factor expression, and ERbeta was not detected. A receiver operating characteristic curve for topoisomerase IIalpha disclosed that above a labeling index of 13, the test had 88% sensitivity and 100% specificity for predicting malignancy. CONCLUSION: Cellular proliferation measured with topoisomerase IIalpha is a simple prognostic marker for malignancy in EPTs, unlike apoptosis, angiogenesis, or the presence of ERbeta, which were not associated with malignant behavior. These findings designate a defined field for future research on endocrine pancreatic carcinogenesis and a possible target for chemotherapeutic agents.

Biochemical characteristics of the gamma-aminobutyric acid system in the insulinoma cell lines HIT-T15, RIN-m5F, betaTC3, and comparison with rat brain.

BACKGROUND: gamma-aminobutyric acid (GABA) is the most abundant inhibitory neurotransmitter in the mammalian brain. Both GABA and its synthesizing enzyme, L-glutamate decarboxylase (GAD), are also present in the insulin-secreting pancreatic beta cells, in which its physiologic role is unclear. We have studied several aspects of the GABA system in the insulinoma cell lines HIT-T15, RIN-m5F, and betaTC3 in comparison with rat brain tissue. METHODS: Insulinoma cell lines and embryonic rat brain cortex neurons were cultured. GAD activity was determined by a radioenzymatic method and the presence of GAD(67) protein was assessed by immunocytochemistry. Amino acid content and the effect of different conditions on the release of endogenous GABA were measured by HPLC and fluorometric detection after o-phthaldialdehyde derivatization. [3H]GABA was used for measuring the uptake of the amino acid in the insulinoma cultures and in rat forebrain synaptosomes. RESULTS: The three insulinoma lines possess GABA and GAD activity at levels of approximately 20% compared with adult rat brain cortex. Dissimilar from the latter, in insulinoma cultures enzyme activity was not enhanced by addition of an excess of the coenzyme pyridoxal-5'-phosphate. Immunocytochemical visualization of GAD showed that the cells in both neuronal cultures and insulinoma lines were GAD(67)-positive, similar to Purkinje cell somata of adult rat cerebellar cortex. [3H]GABA uptake in the cell lines was approximately 10% of that in rat forebrain synaptosomes and showed less ionic and temperature dependence. In both cultured cerebral neurons and RINm5F cells, the addition of arginine induced the release of GABA, whereas neither high K(+) concentration nor glucose had any effect. CONCLUSIONS: The insulinoma cell lines studied possess the same GAD(67) form of the enzyme present in brain. RIN line cells are capable of transporting glutamate. In these cells as well as in cultured cortical neurons, arginine stimulates the release of GABA and glutamate probably as the result of its electrogenic transport. Insulinoma cell lines may therefore be useful to study GABA metabolism and function in pancreatic beta cells.

Insulinoma: diagnostic strategies and surgical treatment. A 22-year experience.

BACKGROUND/AIMS: The efficacy of preoperative localization methods and the results of the surgical treatment of insulinoma were studied. METHODOLOGY: Fifty-nine patients referred for surgical treatment were studied and the results of the diagnostic tools for tumor localization were compared with findings at surgical intervention. The influence of the type of surgical procedure in the immediate and late postoperative course was also studied. RESULTS: Ultrasonography had a sensitivity of 30%, computed tomography 25%, angiography 54%, portal vein sampling 94%, endoscopic ultrasonography 27% and magnetic resonance 17%. Intraoperative palpation localized 98.2% of the tumors and by the addition of intraoperative echography, all lesions were identified. In 55 patients with benign lesions, 22 enucleations, 25 distal pancreatectomies, 7 pancreatectomies plus enucleation and one duodenopancreatectomy were performed. Malignant tumors were treated by pancreatic resection, postoperative hepatic artery embolization and systemic chemotherapy. There was no postoperative mortality. Pancreatic fistula was the most common complication. Three patients who underwent distal pancreatectomy developed late diabetes (9.3%). CONCLUSIONS: Extensive preoperative investigation, mainly with invasive methods, is not indicated and by combining intraoperative palpation and echography most of the cases can be adequately dealt with. Preservation of pancreatic tissue with enucleation and preservation of the spleen are the best choice for treatment of benign insulinomas.