Actinomycin D Arrests Cell Cycle of Hepatocellular Carcinoma Cell Lines and Induces p53-Dependent Cell Death: A Study of the Molecular Mechanism Involved in the Protective Effect of IRS-4.

Actinomycin D (ActD) is an FDA-approved NCI oncology drug that specifically targets and downregulates stem cell transcription factors, which leads to a depletion of stem cells within the tumor bulk. Recently, our research group demonstrated the importance of IRS-4 in the development of liver cancer. In this study, we evaluated the protective effects of IRS-4 against ActD. For this study, three hepatocellular carcinoma cell lines (HepG2, Huh7, and Chang cells) were used to study the mechanism of actinomycin D. Most assays were carried out in the Hep G2 cell line, due to the high expression of stem cell biomarkers. We found that ActD caused HepG2 cell necroptosis characterized by DNA fragmentation, decreased mitochondrial membrane potential, cytochrome c depletion, and decreased the levels of reduced glutathione. However, we did not observe a clear increase in apoptosis markers such as annexin V presence, caspase 3 activation, or PARP fragmentation. ActD produced an activation of MAP kinases (ERK, p38, and JNK) and AKT. ActD-induced activation of AKT and MAP kinases produced an activation of the Rb-E2F cascade together with a blockage of cell cycle transitions, due to c-jun depletion. ActD led to the inhibition of pCdK1 and pH3 along with DNA fragmentation resulting in cell cycle arrest and the subsequent activation of p53-dependent cell death in the HepG2 cell line. Only JNK and AKT inhibitors were protective against the effects of ActD. N-Acetyl-L-cysteine also had a protective effect as it restored GSH levels. A likely mechanism for this is IRS-4 stimulating GCL-GSH and inhibiting the Brk-CHK1-p53 pathway. The assessment of the IRS-4 in cancer biopsies could be of interest to carry out a personalized treatment with ActD.

Possible Role of IRS-4 in the Origin of Multifocal Hepatocellular Carcinoma.

New evidence suggests that insulin receptor substrate 4 (IRS-4) may play an important role in the promotion of tumoral growth. In this investigation, we have evaluated the role of IRS-4 in a pilot study performed on patients with liver cancer. We used immunohistochemistry to examine IRS-4 expression in biopsies of tumoral tissue from a cohort of 31 patient suffering of hepatocellular carcinoma (HCC). We simultaneously analyzed the expression of the cancer biomarkers PCNA, Ki-67, and pH3 in the same tissue samples. The in vitro analysis was conducted by studying the behavior of HepG2 cells following IRS-4 overexpression/silencing. IRS-4 was expressed mainly in the nuclei of tumoral cells from HCC patients. In contrast, in healthy cells involved in portal triads, canaliculi, and parenchymal tissue, IRS-4 was observed in the cytosol and the membrane. Nuclear IRS-4 in the tumoral region was found in 69.9 ± 3.2%, whereas in the surrounding healthy hepatocytes, nuclear IRS-4 was rarely observed. The percentage of tumoral cells that exhibited nuclear PCNA and Ki-67 were 52.1 ± 7%, 6.1 ± 1.1% and 1.3 ± 0.2%, respectively. Furthermore, we observed a significant positive linear correlation between nuclear IRS-4 and PCNA (r = 0.989; p < 0.001). However, when we correlated the nuclear expression of IRS-4 and Ki-67, we observed a significant positive curvilinear correlation (r = 0.758; p < 0.010). This allowed us to define two populations, (IRS-4 + Ki-67 ≤ 69%) and (IRS-4 + Ki-67 > 70%). The population with lower levels of IRS-4 and Ki-67 had a higher risk of suffering from multifocal liver cancer (OR = 16.66; CI = 1.68-164.8 (95%); p < 0.05). Immunoblot analyses showed that IRS-4 in normal human liver biopsies was lower than in HepG2, Huh7, and Chang cells. Treatment of HepG2 with IGF-1 and EGF induced IRS-4 translocation to the nucleus. Regulation of IRS-4 levels via HepG2 transfection experiments revealed the protein's role in proliferation, cell migration, and cell-collagen adhesion. Nuclear IRS-4 is increased in the tumoral region of HCC. IRS-4 and Ki-67 levels are significantly correlated with the presence of multifocal HCC. Moreover, upregulation of IRS-4 in HepG2 cells induced proliferation by a ß-catenin/Rb/cyclin D mechanism, whereas downregulation of IRS-4 caused a loss in cellular polarity and in its adherence to collagen as well as a gain in migratory and invasive capacities, probably via an integrin α2 and focal adhesion cascade (FAK) mechanism.

Nutritional Components in Western Diet Versus Mediterranean Diet at the Gut Microbiota-Immune System Interplay. Implications for Health and Disease.

The most prevalent diseases of our time, non-communicable diseases (NCDs) (including obesity, type 2 diabetes, cardiovascular diseases and some types of cancer) are rising worldwide. All of them share the condition of an "inflammatory disorder", with impaired immune functions frequently caused or accompanied by alterations in gut microbiota. These multifactorial maladies also have in common malnutrition related to physiopathology. In this context, diet is the greatest modulator of immune system-microbiota crosstalk, and much interest, and new challenges, are arising in the area of precision nutrition as a way towards treatment and prevention. It is a fact that the westernized diet (WD) is partly responsible for the increased prevalence of NCDs, negatively affecting both gut microbiota and the immune system. Conversely, other nutritional approaches, such as Mediterranean diet (MD), positively influence immune system and gut microbiota, and is proposed not only as a potential tool in the clinical management of different disease conditions, but also for prevention and health promotion globally. Thus, the purpose of this review is to determine the regulatory role of nutritional components of WD and MD in the gut microbiota and immune system interplay, in order to understand, and create awareness of, the influence of diet over both key components.

Desmoid-type Fibromatosis of the Splenic Hilum: A Rare Tumor at an Unusual Location.

Desmoid-type fibromatoses (DTFs), also known as desmoid tumors, are benign but infiltrative neoplasms that often appear next to previous surgical site. Intra-abdominal tumors usually involve the mesentery, but splenic hilum is an unusual localization. We present a case of a desmoid tumor of the splenic hilum laparoscopically resected in a 70-year-old male with a previous history of chromophobe renal cell carcinoma and ocular spindle melanoma. Although benign, desmoid tumors might be infiltrative and produce serious complications. Treatment remains controversial, ranging from surgery and medical therapies to observation. Management of DTF must be individualized, considering the risk of complications and recurrence.

Duodenal Rupture after Blunt Abdominal Trauma by Bicycle Handlebar: Case Report and Literature Review.

Blunt abdominal trauma is most frequent in the pediatric population. Duodenal lesions after abdominal trauma in children are infrequent and tend to be secondary to traffic accidents. It is up to five times more frequent in males, with an average age between 16 and 30 years. Bicycle accidents continue to lead to morbidity and mortality in children, representing between 5% and 14% of total blunt abdominal injuries. The diagnosis of duodenal injuries after trauma is difficult and requires a high index of clinical suspicion. We present the case of a 17-year-old patient seen in the emergency room after falling off his bicycle and presented a blunt trauma in the epigastric region. On physical examination, there was a swelling in the upper right abdominal quadrant and epigastrium with tenderness on deep palpation. He presented with hematemesis without hemodynamic repercussion. A contrast abdominal computed tomography was performed and he was diagnosed with third-part duodenal rupture. A resection of the perforated third-part duodenal rupture was performed, and the transit was reconstructed using a Roux-Y duodenojejunostomy. The postoperative period was uneventful and the patient was discharged after 16 days of stay. Duodenal injury is very rare, produced by high-energy trauma. They rarely present as single lesions as other visceral lesions are usually associated. The early diagnosis is important to reduce the morbidity and mortality.

Septic thrombosis of internal jugular vein: a rare cause of uncontrollable hiccups.

Septic thrombosis of the internal jugular vein is a possible complication related to central venous catheters. The enlargement of the diameter of the jugular vein can stimulate phrenic nerve causing hiccups and, septic thrombus can metastasize to different organs threating patient's life. Diagnosis of septic thrombosis of internal jugular vein should be confirmed with a cervicothoracic CT-scan. Its management consists of catheter's removal, antibiotic treatment and anticoagulation in high-risk patients. Surgical intervention might be considered if conservative treatment fails.

Primary Hyperparathyroidism due to Parathyroid Adenoma Originated from Supernumerary Gland.

INTRODUCTION: The variability of the location of the parathyroid glands is directly related to the events that occur during embryonic development. The impact that an individual submits more than four parathyroid glands is close to 13%. However the presentation of a parathyroid adenoma in a supernumerary gland is an uncommon event. CASE REPORT: A 30-year-old man diagnosed with primary hyperparathyroidism with matching findings on ultrasonography and scintigraphy for parathyroid adenoma localization lower left regarding the thyroid gland. A cervicotomy explorer showed four orthotopic parathyroid glands. The biopsy of the inferior left gland was normal. No signs of adenoma were seen in the biopsy. Following mobilization of the ipsilateral thyroid lobe, fifth parathyroid gland was found increased significantly in size than proceeded to remove, confirming the diagnosis of adenoma. After the excision, the levels of serum calcium and parathyroid hormone were normalized. CONCLUSIONS: The presentation of a parathyroid adenoma in a supernumerary gland is a challenge for the surgeon. The high sensitivity having different imaging techniques has been a key to locate preoperatively the pathological parathyroid gland. Analytical or clinical persistence of primary hyperparathyroidism after parathyroid surgery can occur if the location of the adenoma is a supernumerary or ectopic gland location.

Insulin receptor substrate-4 is overexpressed in colorectal cancer and promotes retinoblastoma-cyclin-dependent kinase activation.

BACKGROUND: Insulin receptor substrate 4 (IRS-4) is an adaptor protein for which new evidence suggests plays a role in tumour promotion. METHODS: We described nuclear IRS-4 in RKO colon cancer cell lines in biopsies of patients with colorectal cancer (CRC) (n = 20) and in matched adjacent normal colorectal (MANC) tissue (n = 20). RESULTS: Treatment with physiological doses of IGF-1 promoted nuclear influx of IRS-4 from cellular cytosol in RKO cells. When exogenous IRS-4 was overexpressed in RKO cells, there was an increase in cyclin D1, cyclin E, E2F1, pRB Ser 809/811 and pRB Ser 705 levels compared with the empty vector-transfected cells. Some of these changes returned to control values after wortmannin treatment. Subcellular fractionation showed an overexpression of IRS-4 in the cytoplasm, membrane, and nuclei of tumour samples, whereas the levels of the protein were barely detectable in the three compartments of normal samples. Immunohistochemical studies showed positive nuclear IRS-4 staining in over 74% of the tumour cells. IRS-4 was strongly overexpressed in tumoural tissues from CRC patients compared to MANC tissues. The up-regulation of IRS-4 in CRC samples correlated significantly with the increase of several G1 checkpoint proteins including cyclin D1 (r = 0.6662), Rb (r = 0.7779), pRb Serine 809/811 (r = 0.6864), pRb serine 705 (r = 0.6261) and E2F1 (r = 0.8702). CONCLUSIONS: Taken together, our findings suggest that IRS-4 promotes retinoblastoma-cyclin-dependent kinase activation and it may serve as a pharmacological target since its expression is very low in normal tissue, including colonic epithelium.

Overexpression of insulin receptor substrate-4 is correlated with clinical staging in colorectal cancer patients.

Insulin receptor substrate-4 (IRS-4), a poorly studied member of the IRS family, may play an important role in colorectal cancer (CRC) tumourigenesis. The aim of this pilot study was to elucidate the potential role of IRS-4 in colorectal carcinogenesis by evaluating IRS-4 expression in different types of colorectal tumours (n = 20) and comparing its expression to normal mucosa (n = 20). Tissue samples were collected from 18 patients with CRC and 2 with precancerous lesions (tubulovillous adenomas), all of whom were undergoing potentially curative surgery. IRS-4 expression was evaluated using immunohistochemical staining and compared to clinicopathological features. In normal colonic crypts, the subcellular localization of IRS-4 varied from the crypt base compartment to the surface epithelium. Nuclear IRS-4 staining decreased while non-nuclear IRS-4 increased as cells approached the top of the crypt. In the patients studied, colorectal tumours showed a significant (p < 0.001) increase of IRS-4 expression compared with adjacent normal tissue. Furthermore, nuclear IRS-4 intensity of CRC patients was significantly (p < 0.0001) higher in colonic tumoural tissue than in paired normal specimens. Tumour expression of IRS-4 in CRC patients was positively associated with T (p < 0.0001) and N (p < 0.05), of TNM (tumour and nodes and metastasis) staging system. Taken together, these results suggest that increase of IRS-4 expression may be involved to some extent in colorectal cancer.

Ileocolic intussusception due to giant ileal lipoma: Review of literature and report of a case.

INTRODUCTION: Intussusception in adults accounts for less than 5% of all intussusceptions. It occurs when a segment of intestine invaginates into itself. PRESENTATION OF A CASE: We report a case of ileocolic intussusception in an adult caused by a giant ileal lipoma. DISCUSSION: Intussusceptions can be classified as ileocolic, ileocecal, colo-colic and ileo-ileal. Most are due to neoplasms (60% malign and 24-40% benign). In the colon, the possibility of malignancy is higher than in small intestine. Lipomas are the most common benign mesenchymal intestinal tumors, accounting for less than 5% of all gastrointestinal tumors. They are more frequent in colon than small intestine. Small lipomas (less than 2cm) are usually asymptomatic. Larger lesions may produce symptoms such as abdominal pain, obstruction or intussusception. Lipomas can be diagnosed with endoscopy, capsule endoscopy, barium enemas, CT and US. CONCLUSION: Intussusceptions in adults is a rare condition, most of them are caused by a malign neoplasms followed by benign neoplasms. US and CT are useful for diagnosis. Surgery is mandatory.