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Background: Colorectal cancer (CRC) is characterized by a pro-inflammatory microenvironment and features high-energy-supply molecules that assure tumor growth. A still less studied macromolecule is inorganic polyphosphate (iPolyP), a high-energy linear polymer that is ubiquitous in all forms of life. Made up of hundreds of repeated orthophosphate units, iPolyP is essential for a wide variety of functions in mammalian cells, including the regulation of proliferative signaling pathways. Some evidence has suggested its involvement in carcinogenesis, although more studies need to be pursued. Moreover, iPolyP regulates several homeostatic processes in animals, spanning from energy metabolism to blood coagulation and tissue regeneration., Results: In this study, we tested the role of iPolyP on CRC proliferation, using in vitro and ex vivo approaches, in order to evaluate its effect on tumor growth. We found that iPolyP is significantly increased in tumor tissues, derived from affected individuals enrolled in this study, compared to the corresponding peritumoral counterparts. In addition, iPolyP signaling occurs through the TRPM8 receptor, a well-characterized Na + and Ca 2+ ion channel often overexpressed in CRC and linked with poor prognosis, thus promoting CRC cell proliferation. The pharmacological inhibition of TRPM8 or RNA interference experiments performed in established CRC cell lines, such as Caco-2 and SW620, showed that the involvement of TRPM8 is essential, greater than that of the other two known iPolyP receptors, P2Y1 and RAGE. The presence of iPolyP drives cancer cells towards the mitotic phase of the cell cycle by enhancing the expression of ccnb1 , which encodes the Cyclin B protein. In vitro 2D and 3D data reflected the ex vivo results, obtained by the generation of CRC-derived organoids, which increased in size., Conclusions: These results indicate that iPolyP may be considered a novel and unexpected early biomarker supporting colorectal cancer cell proliferation.

Over 40,000 patients in the United States are estimated to suffer from end-stage liver disease and acute hepatic failure, for which liver transplantation is the only available therapy. Human primary hepatocytes (HPH) have not been employed as a therapeutic tool due to the difficulty in growing and expanding them in vitro, their sensitivity to cold temperatures, and tendency to dedifferentiate following two-dimensional culture. The differentiation of human-induced pluripotent stem cells (hiPSCs) into liver organoids (LO) has emerged as a potential alternative to orthotropic liver transplantation (OLT). However, several factors limit the efficiency of liver differentiation from hiPSCs, including a low proportion of differentiated cells capable of reaching a mature phenotype, the poor reproducibility of existing differentiation protocols, and insufficient long-term viability in vitro and in vivo. This review will analyze various methodologies being developed to improve hepatic differentiation from hiPSCs into liver organoids, paying particular attention to the use of endothelial cells as supportive cells for their further maturation. Here, we demonstrate why differentiated liver organoids can be used as a research tool for drug testing and disease modeling, or employed as a bridge for liver transplantation following liver failure.

One of the main causes of post-transplant-associated morbidity and mortality is cancer. The aims of the project were to study the neoplastic risk within the kidney transplant population and identify the determinants of this risk. A cohort of 462 renal transplant patients from 2010 to 2020 was considered. The expected incidence rates of post-transplant cancer development in the referenced population, the standardized incidence ratios (SIR) taking the Italian population as a comparison, and the absolute risk and the attributable fraction were extrapolated from these cohorts of patients. Kidney transplant recipients had an overall cancer risk of approximately three times that of the local population (SIR 2.8). A significantly increased number of cases were observed for Kaposi's sarcoma (KS) (SIR 195) and hematological cancers (SIR 6.8). In the first 3 years post-transplant, the risk to develop either KS or hematological cancers was four times higher than in the following years; in all cases of KS, the diagnosis was within 2 years from the transplant. Post-transplant immunosuppression represents the cause of 99% of cases of KS and 85% of cases of lymphomas, while only 39% is represented by solid tumors. Data related to the incidence, the percentages attributable to post-transplant immunosuppression, and the time of onset of neoplasms, particularly for KS and hematological tumors could help improve the management for the follow-up in these patients., (© 2023. The Author(s).)

Objectives: The following retrospective study was devised with the aim of evaluating the correlation between OSAS and various anatomical factors., Material and Methods: Thirty-seven patients over the age of 40 were analyzed, of which 19 were classified as OSAS cases and 18 as control cases. For each, 17 anatomical variables were identified and examined using Invivo Dental software on CBCT scans, WebCeph software on laterolateral teleradiographs, and Rhinoceros 6.0 software on dental casts., Results: A generalized linear model of all the anatomical factors identified only two statistically significant variables. Specifically, the total volume of the palate displayed a inverse correlation with OSAS, while the distance between the S point and the Go point (S-Go) exhibited a direct correlation with the disease., Conclusion: The likelihood of an individual having OSAS appears to decrease as the volume of the palate increases but increase as the lingual measure S-Go increases.

Liver involvement after abdominal blunt trauma must be expected, and in up to 30% of cases, spleen, kidney, and pancreas injuries may coexist. Whenever hemodynamics conditions do not contraindicate the overcoming of the ancient dogma according to which exploratory laparotomy should be performed after every major abdominal trauma, a CT scan has to clarify the liver lesions so as to determine the optimal management strategy. Except for complete vascular avulsion, no liver trauma grade precludes nonoperative management. Every attempt to treat the injured liver by avoiding a strong surgical approach may be considered. Each time, a nonoperative management (NOM) consisting of a basic "wait and see" attitude combined with systemic support and blood replacement are inadequate. Embolization should be considered to stop the bleeding. Percutaneous drainage of collections, endoscopic retrograde cholangiopancreatography (ERCP) with papilla sphincterotomy or stent placement and percutaneous transhepatic biliary drainage (PTBD) may avoid, or at least delay, surgical reconstruction or resection until systemic and hepatic inflammatory remodeling are resolved. The pathophysiological principle sustaining these leanings is based on the opportunity to limit the further release of cell debris fragments acting as damage-associated molecular patterns (DAMPs) and the following stress response associated with the consequent immune suppression after trauma. The main goal will be a faster recovery combined with limited cell death of the liver through the ischemic events that may directly follow the trauma, exacerbated by hemostatic procedures and surgery, in order to reduce the gross distortion of a regenerated liver.

Human-induced pluripotent stem cells (hiPSCs) constitute a great source to generate specialized cells that can be employed in cell replacement therapy for a number of degenerative diseases. In this chapter, I describe a strategy to mass-produce fully functional hepatocyte organoids using hiPSCs interlaced with human adipose microvascular endothelial cells (HAMEC). Our unique technology employs a two-step strategy, consisting of the scalable generation of nearly spherical uniform-sized human embryoid bodies (hEBs), and the subsequent employment of a four-step hepatocyte differentiation approach for the generation of hepatocyte organoids that display all the characteristics of human primary hepatocytes. In this chapter, we also describe methodologies to characterize the hepatocyte organoids by using different biomolecular assays., (© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Human induced pluripotent stem cells (hiPSCs) represent a powerful tool for the generation of specialized cells to be used in regenerative medicine as well as hepatocellular repopulation tool to treat liver metabolic diseases such as nonalcoholic steatohepatitis (NASH). Here we describe a strategy to obtain fully functional liver organoids from hiPSCs in a scalable manner. Our approach uses a two-step process, with a first step involving the scalable formation of homogeneous and uniform-sized human embryoid bodies (hEBs), followed by the application of a four-step liver differentiation protocol for the derivation of liver organoids that possess all the features of primary human hepatocytes. This chapter will also illustrate the characterization of the liver organoids by directed biomolecular techniques., (© 2022. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

A laparoscopic approach is suggested with the highest grade of recommendation for acute cholecystitis, perforated gastroduodenal ulcers, acute appendicitis, gynaecological disorders, and non-specific abdominal pain (NSAP). To date, the main qualities of laparoscopy for these acute surgical scenarios are clearly stated: quicker surgery, faster recovery and shorter hospital stay. For the remaining surgical emergencies, as well as for abdominal trauma, the role of laparoscopy is still a matter of debate. Patients might benefit from a laparoscopic approach only if performed by experienced teams and surgeons which guarantee a high standard of care. More precisely, laparoscopy can limit damage to the tissue and could be effective for the reduction of the overall amount of cell debris, which is a result of the intensity with which the immune system reacts to the injury and the following symptomatology. In fact, these fragments act as damage-associated molecular patterns (DAMPs). DAMPs, as well as pathogen associated molecular patterns (PAMPs), are recognised by both surface and intracellular receptors of the immune cells and activate the cascade which, in critically ill surgical patients, is responsible for a deranged response. This may result in the development of progressive and multiple organ dysfunctions, manifesting with acute respiratory distress syndrome (ARDS), coagulopathy, liver dysfunction and renal failure. In conclusion, none of the emergency surgical scenarios preclude laparoscopy, provided that the surgical tactic could ensure sufficient cleaning of the abdomen in addition to resolving the initial tissue damage caused by the "trauma".

Organoids formed from human induced pluripotent stem cells (hiPSCs) could be a limitless source of functional tissue for transplantations in many organs. Unfortunately, fine-tuning differentiation protocols to form large quantities of hiPSC organoids in a controlled, scalable, and reproducible manner is quite difficult and often takes a very long time. Recently, we introduced a new approach of rapid organoid formation from dissociated hiPSCs and endothelial cells using microfabricated cell-repellent microwell arrays. This approach, when combined with real-time label-free Raman spectroscopy of biochemical composition changes and confocal light scattering spectroscopic microscopy of chromatin transition, allows for monitoring live differentiating organoids without the need to sacrifice a sample, substantially shortening the time of protocol fine-tuning. We used this approach to both culture and monitor homogeneous liver organoids that have the main functional features of the human liver and which could be used for cell transplantation liver therapy in humans., (Copyright © 2021 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works. Distributed under a Creative Commons Attribution NonCommercial License 4.0 (CC BY-NC).)

Despite advances in stem cell research, cell transplantation therapy for liver failure is impeded by a shortage of human primary hepatocytes (HPH), along with current differentiation protocol limitations. Several studies have examined the concept of co-culture of human induced pluripotent cells (hiPSCs) with various types of supporting non-parenchymal cells to attain a higher differentiation yield and to improve hepatocyte-like cell functions both in vitro and in vivo. Co-culturing hiPSCs with human endothelial cells (hECs) is a relatively new technique that requires more detailed studies. Using our 3D human embryoid bodies (hEBs) formation technology, we interlaced Human Adipose Microvascular Endothelial Cells (HAMEC) with hiPSCs, leading to a higher differentiation yield and notable improvements across a wide range of hepatic functions. We conducted a comprehensive gene and protein secretion analysis of our HLCs coagulation factors profile, showing promising results in comparison with HPH. Furthermore, a stage-specific glycomic analysis revealed that the differentiated hepatocyte-like clusters (HLCs) resemble the glycan features of a mature tissue rather than cells in culture. We tested our HLCs in animal models, where the presence of HAMEC in the clusters showed a consistently better performance compared to the hiPSCs only group in regard to persistent albumin secretion post-transplantation.

Intestinal injury and immune dysfunction are commonly encountered after irradiation therapy. While the curative abilities of ginseng root have been reported in prior studies, there is little known regarding its role in immunoregulation of intestinal repairability in cancer patients treated with irradiation. Our current study aims to closely examine the protective effects of ginseng-derived small molecule oligopeptides (Panax ginseng C. A. Mey.) (GOP) against irradiation-induced immune dysfunction and subsequent intestinal injury, using in vitro and in vivo models. Expectedly, irradiation treatment resulted in increased intestinal permeability along with mucosal injury in both Caco-2 cells and mice, probably due to disruption of the intestinal epithelial barrier, leading to high plasma lipopolysaccharide (LPS) and pro-inflammatory cytokines levels. However, when the cells were treated with GOP, this led to diminished concentration of plasma LPS and cytokines (IL-1 and TNF-α), suggesting its dampening effect on inflammatory and oxidative stress, and potential role in restoring normal baseline intestinal permeability. Moreover, the Caco-2 cells treated with GOP showed high trans-epithelial electrical resistance (TEER) and low FITC-dextran paracellular permeability when compared to the control group. This could be explained by the higher levels of tight junction proteins (ZO-1 and Occludin) expression along with reduced expression of the apoptosis-related proteins (Bax and Caspase-3) noticed in the GOP-treated cells, highlighting its role in preserving intestinal permeability, through prevention of their degradation while maintaining normal levels of expression. Further confirmatory in vivo data showed that GOP-treated mice exhibited high concentrations of lymphocytes (CD3 + , CD4 + , CD8 + ) in the intestine, to rescue the irradiation-induced damage and restore baseline intestinal integrity. Therefore, we propose that GOP can be used as an adjuvant therapy to attenuate irradiation-induced immune dysfunction and intestinal injury in cancer patients.

The discovery of human induced pluripotent stem cells (hiPSCs) is a promising advancement in the field of regenerative and personalized medicine. Expression of SOX2, KLF4, OCT4 and MYC transcription factors induces the nuclear reprogramming of somatic cells into hiPSCs that share striking similarities with human embryonic stem cells (hESCs). However, several studies have demonstrated that hESCs and hiPSCs could lead to teratoma formation in vivo, thus limiting their current clinical applications. Aberrant cell cycle regulation of hESCs is linked to centrosome amplification, which may account, for their enhanced chromosomal instability (CIN), and thus increase their tumorigenicity. Significantly, the tumor suppressor p53 plays a key role as a 'guardian of reprogramming', safeguarding genomic integrity during hiPSC reprogramming. Nevertheless, the molecular mechanisms leading to development of CIN during reprogramming and increased tumorigenic potential of hiPSCs remains to be fully elucidated. In the present study, we analyzed CIN in hiPSCs derived from keratinocytes and established that chromosomal and mitotic aberrations were linked to centrosome amplification, Aurora-A overexpression, abrogation of p53-mediated G1/S cell cycle checkpoint and loss of Rb tumor-suppressor function. When hiPSCs were transplanted into the kidney capsules of immunocompromised mice, they developed high-grade teratomas characterized by the presence of cells that exhibited non-uniform shapes and sizes, high nuclear pleomorphism and centrosome amplification. Significantly, ex vivo cells derived from teratomas exhibited high self-renewal capacity that was linked to Aurora-A kinase activity and gave rise to lung metastasis when injected into the tail vein of immunocompromised mice. Collectively, these findings demonstrated a high risk for malignancy of hiPSCs that exhibit Aurora-A overexpression, loss of Rb function, centrosome amplification and CIN. Based on these findings, we proposed that Aurora-A-targeted therapy could represent a promising prophylactic therapeutic strategy to decrease the likelihood of CIN and development of aggressive teratomas derived from hiPSCs.

Traditionally used as a restorative medicine, ginseng (Panax ginseng Meyer) has been the most widely used and acclaimed herb in Chinese communities for thousands of years. To investigate the immune-modulating activity of ginseng oligopeptides (GOP), 420 healthy female BALB/c mice were intragastrically administered distilled water (control), whey protein (0.15 g per kg body weight (BW)), and GOP 0.0375, 0.075, 0.15, 0.3 and 0.6 g per kg BW for 30 days. Blood samples from mice were collected from the ophthalmic venous plexus and then sacrificed by cervical dislocation. Seven assays were conducted to determine the immunomodulatory effects of GOP on innate and adaptive immune responses, followed by flow cytometry to investigate spleen T lymphocyte sub-populations, multiplex sandwich immunoassays to investigate serum cytokine and immunoglobulin levels, and ELISA to investigate intestinally secreted immunoglobulin to study the mechanism of GOP affecting the immune system. Our results showed that GOP was able to enhance innate and adaptive immune responses in mice by improving cell-mediated and humoral immunity, macrophage phagocytosis capacity and NK cell activity. Notably, the use of GOP revealed a better immune-modulating activity compared to whey protein. We conclude that the immune-modulating activity might be due to the increased macrophage phagocytosis capacity and NK cell activity, and the enhancement of T and Th cells, as well as IL-2, IL-6 and IL-12 secretion and IgA, IgG1 and IgG2b production. These results indicate that GOP could be considered a good candidate that may improve immune functions if used as a dietary supplement, with a dosage that ranges from 0.3 to 0.6 g per kg BW.

The sensitivity of lymph node core-needle biopsy under imaging guidance requires validation. We employed power Doppler ultrasonography (PDUS) to select the lymph node most suspected of malignancy and to histologically characterize it through the use of large cutting needle. Institutional review board approval and informed consent were obtained for this randomized clinical trial. In a single center between 1 January 2009 and 31 December 2015, patients with lymph node enlargement suspected for lymphoma were randomly assigned (1:1) to biopsy with either standard surgery or PDUS-guided 16-gauge modified Menghini needle. The primary endpoint was the superiority of sensitivity for the diagnosis of malignancy for core-needle cutting biopsy (CNCB). Secondary endpoints were times to biopsy, complications, and costs. A total of 376 patients were randomized into the two arms and received allocated biopsy. However, four patients undergoing CNCB were excluded for inadequate samples; thus, 372 patients were analyzed. Sensitivity for the detection of malignancy was significantly better for PDUS-guided CNCB [98.8%; 95% confidence interval (CI), 95.9-99.9] than standard biopsy (88.7%; 95% CI, 82.9-93; P < 0.001). For all secondary endpoints, the comparison was significantly disadvantageous for conventional approach. In particular, estimated cost per biopsy performed with standard surgery was 24-fold higher compared with that performed with CNCB. The presence of satellite enlarged reactive and/or necrotic lymph nodes may impair the success of an open surgical biopsy (OSB). PDUS and CNCB with adequate gauge are diagnostic tools that enable effective, safe, fast, and low-cost routine biopsy for patients with suspected lymphoma, avoiding psychological and physical pain of an unnecessary surgical intervention.

The aim of the study was to assess incidence rate, hormonal activity, and local invasiveness and evaluate outcomes of so-diagnosed atypical pituitary adenomas that underwent endoscopic endonasal surgery at the Division of Neurosurgery of Università degli Studi di Napoli Federico II. According to the 2004 WHO classification, atypical pituitary adenomas are defined by an invasive growth, Ki-67/MIB-1 proliferative index greater than 3 %, high p53 immunoreactivity, and increased mitotic activity. A retrospective analysis of a series of 434 pituitary adenomas that underwent endoscopic endonasal surgery at our department between March 2007 and February 2013 was performed. Fifty adenomas (11.5 %) met the criteria of diagnosis of atypical lesions; 10 (21.6 %) of the 50 patients were recurrent tumors with a previous transsphenoidal surgery. Forty-one (82 %) were macroadenomas, and 21/50 (42 %) showed a clear invasion of the cavernous sinus. Histotype of atypical adenomas figured out to be nonfunctioning in 23 cases (46 %), PRL secreting in 10 cases (20 %), ACTH secreting and GH secreting each apart in 8 patients (16 %), and in a single case a GH/PRL secreting adenoma (2 %). The Ki-67 labeling index ranged from 3.5 to 22.5 % (mean 5.6 %). Tumor recurrence was observed in six cases (12 %) after a mean time of 18 months (range 9-24 months). Mean follow-up was 36.5 months (range 2-80 months). Atypical pituitary adenomas account for ca. 10 % of all pituitary adenomas; these lesions have peculiar features. It should be considered that a strong immunopositivity of p53 and higher Ki-67 LI could predict an increased risk of tumor recurrence, but more studies and larger series are expected to confirm and enlarge the diagnostic and therapeutic management process of these lesions.

Treatment of acute liver failure by cell transplantation is hindered by a shortage of human hepatocytes. Current protocols for hepatic differentiation of human induced pluripotent stem cells (hiPSCs) result in low yields, cellular heterogeneity, and limited scalability. In the present study, we have developed a novel multicellular spheroid-based hepatic differentiation protocol starting from embryoid bodies of hiPSCs (hiPSC-EBs) for robust mass production of human hepatocyte-like cells (HLCs) using two novel inhibitors of the Wnt pathway. The resultant hiPSC-EB-HLCs expressed liver-specific genes, secreted hepatic proteins such as Albumin, Alpha Fetoprotein, and Fibrinogen, metabolized ammonia, and displayed cytochrome P450 activities and functional activities typical of mature primary hepatocytes, such as LDL storage and uptake, ICG uptake and release, and glycogen storage. Cell transplantation of hiPSC-EB-HLC in a rat model of acute liver failure significantly prolonged the mean survival time and resolved the liver injury when compared to the no-transplantation control animals. The transplanted hiPSC-EB-HLCs secreted human albumin into the host plasma throughout the examination period (2 weeks). Transplantation successfully bridged the animals through the critical period for survival after acute liver failure, providing promising clues of integration and full in vivo functionality of these cells after treatment with WIF-1 and DKK-1.

Sclerostin is a marker of low-turnover bone disease in end stage renal disease patients. The aim of this study was to evaluate serum sclerostin in uremic patients, analyzing its behavior during a single hemodialysis session. Twenty-one adult patients on intermittent hemodialysis treatment were enrolled. Acetate Free Bio-filtration (AFB) was the technique employed. Uremic patients were characterized by higher levels of serum sclerostin when compared with values observed in healthy subjects. Sclerostin assessed in pre-dialysis samples was 1.4 ± 1.02 ng/mL, whereas, in post dialysis samples, a reduction of sclerostin values was observed (0.8 ± 0.6 ng/mL; p: 0.008). Sclerostin correlated with parameters of dialysis adequacy, such as creatinine levels and Kt/V values, and it was significantly associated with atherosclerotic disease. Receiver operating characteristics analysis revealed a good diagnostic profile in identifying atherosclerotic disease. Sclerostin, a full dialyzable substance during AFB dialysis, is closely associated with atherosclerotic disease. Its reduction obtained through AFB could represent a defensive mechanism, improving vascular disease and renal osteodystrophy.

Context: Available markers are not reliable parameters to early detect kidney injury in transplanted patients., Objective: Examine neutrophil gelatinase associated lipocalin (NGAL) in early detection of delayed graft function (DGF) and as a long-term predictor of graft outcome., Patients and Methods: NGAL was evaluated in 124 transplanted patients., Results: Urinary NGAL levels were associated to a 10% (HR: 1.10; 95% CI: 1.04-1.25; p < 0.001) and 15% (HR: 1.15; 95% CI: 1.09-1.26; p < 0.001) increased risk of DGF and allograft nephropathy progression, respectively., Conclusion: NGAL reflects the entity of renal impairment in transplanted patients, representing a biomarker and an independent risk factor for DGF and chronic allograft nephropathy progression.

Introduction: Hepatocyte cell transplantation can be life-saving in patients with acute liver failure (ALF); however, primary human hepatocyte transplantation is limited by the scarcity of donor hepatocytes. We investigated the effect of stem cell-derived, hepatocyte-like cells in an animal xenotransplant model of ALF., Methods: Intraperitoneal d-galactosamine was used to develop a lethal model of ALF in the rat. Human induced pluripotent stem cells (iPSC), human mesenchymal stem cells, and human iPSC combined with human endothelial cells (iPSC + EC) were differentiated into hepatocyte-like cells and transplanted into the spleens of athymic nude rats with ALF., Results: A reproducible lethal model of ALF was achieved with nearly 90% death within 3 days. Compared with negative controls, rats transplanted with stem cell-derived, hepatocyte-like cells were associated with increased survival. Human albumin was detected in the rat serum 3 days after transplantation in more than one-half the animals transplanted with hepatocyte-like cells. Only animals transplanted with iPSC + EC-derived hepatocytes had serum human albumin at 14 days posttransplant. Transplanted hepatocyte-like cells homed to the injured rat liver, whereas the ECs were only detected in the spleen., Conclusion: Transplantation of stem cell-derived, hepatocyte-like cells improved survival with evidence of in vivo human albumin production. Combining ECs may prolong cell function after transplantation., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Human pluripotent stem cells (hPSCs) are powerful tools for regenerative therapy and studying human developmental biology, attributing to their ability to differentiate into many functional cell types in the body. The main challenge in realizing hPSC potential is to guide their differentiation in a well-controlled manner. One way to control the cell differentiation process is to recapitulate during in vitro culture the key events in embryogenesis to obtain the three developmental germ layers from which all cell types arise. To achieve this goal, many techniques have been tested to obtain a cellular cluster, an embryoid body (EB), from both mouse and hPSCs. Generation of EBs that are homogeneous in size and shape would allow directed hPSC differentiation into desired cell types in a more synchronous manner and define the roles of cell-cell interaction and spatial organization in lineage specification in a setting similar to in vivo embryonic development. However, previous success in uniform EB formation from mouse PSCs cannot be extrapolated to hPSCs possibly due to the destabilization of adherens junctions on cell surfaces during the dissociation into single cells, making hPSCs extremely vulnerable to cell death. Recently, new advances have emerged to form uniform human embryoid bodies (hEBs) from dissociated single cells of hPSCs. In this review, the existing methods for hEB production from hPSCs and the results on the downstream differentiation of the hEBs are described with emphases on the efficiency, homogeneity, scalability, and reproducibility of the hEB formation process and the yield in terminal differentiation. New trends in hEB production and directed differentiation are discussed.

Myelolipomas are uncommon benign tumors composed of mature adipose tissue mixed with hematopoietic elements; these tumors can occur in both the adrenal glands and extra-adrenal locations, the presacral region being the most frequent extra-adrenal site. We present a case of presacral myelolipoma diagnosed by fine needle aspiration (FNA) and core needle biopsy (CNB) in a 55-year-old woman with concurrent invasive ductal breast cancer. TC and RM imaging were consistent with the diagnosis of presacral myelolipoma. The lesion was discovered incidentally during the staging procedure for breast malignancy. The purpose of our work is to describe the FNA and CNB finding in combination with the imaging features of this uncommon lesion., (Copyright © 2014 Elsevier GmbH. All rights reserved.)

A simple, scalable, and reproducible technology that allows direct formation of large numbers of homogeneous and synchronized embryoid bodies (EBs) of defined sizes from dissociated human induced pluripotent stem cells (hiPSCs) was developed. Non-cell-adhesive hydrogels were used to create round-bottom microwells to host dissociated hiPSCs. No Rho-associated kinase inhibitor (ROCK-i), or centrifugation was needed and the side effects of ROCK-i can be avoided. The key requirement for the successful EB formation in addition to the non-cell-adhesive round-bottom microwells is the input cell density per microwell. Too few or too many cells loaded into the microwells will compromise the EB formation process. In parallel, we have tested our microwell-based system for homogeneous hEB formation from dissociated human embryonic stem cells (hESCs). Successful production of homogeneous hEBs from dissociated hESCs in the absence of ROCK-i and centrifugation was achieved within an optimal range of input cell density per microwell. Both the hiPSC- and hESC-derived hEBs expressed key proteins characteristic of all the three developmental germ layers, confirming their EB identity. This novel EB production technology may represent a versatile platform for the production of homogeneous EBs from dissociated human pluripotent stem cells (hPSCs).

We report a technology to form human embryoid bodies (hEBs) from singularized human embryonic stem cells (hESCs) without the use of the p160 rho-associated coiled-coil kinase inhibitor (ROCKi) or centrifugation (spin). hEB formation was tested under four conditions: +ROCKi/+spin, +ROCKi/-spin, -ROCKi/+spin, and -ROCKi/-spin. Cell suspensions of BG01V/hOG and H9 hESC lines were pipetted into non-adherent hydrogel substrates containing defined microwell arrays. hEBs of consistent size and spherical geometry can be formed in each of the four conditions, including the -ROCKi/-spin condition. The hEBs formed under the -ROCKi/-spin condition differentiated to develop the three embryonic germ layers and tissues derived from each of the germ layers. This simplified hEB production technique offers homogeneity in hEB size and shape to support synchronous differentiation, elimination of the ROCKi xeno-factor and rate-limiting centrifugation treatment, and low-cost scalability, which will directly support automated, large-scale production of hEBs and hESC-derived cells needed for clinical, research, or therapeutic applications.

Purpose: To compare the use of fluorine 18 fluorodeoxyglucose (FDG) positron emission tomography (PET)/computed tomography (CT) with the use of a combination of ultrasonography (US) and chest radiography for systematic follow-up of patients with high-risk Hodgkin lymphoma., Materials and Methods: Institutional review board approval and informed consent were obtained. In a single center between January 2001 and December 2009, patients with advanced-stage Hodgkin lymphoma who had responded completely to first-line treatment were randomly assigned (1:1) to follow-up with either PET/CT or US/chest radiography. Follow-up included clinical and imaging procedures at 4, 8, 12, 16, 20, 24, 30, 36, 48, 60, 84, and 108 months after treatment discontinuation. When clinical and/or imaging results were positive, recurrence was confirmed histologically. The primary endpoint was to compare the sensitivity of the two follow-up imaging approaches. Secondary endpoints were their specificity, positive and negative predictive values, time to recurrence detection, radiation risks, and costs., Results: A total of 300 patients were randomized into the two arms. The study was closed after a median follow-up time of 60 months, with a relapse rate of 27%. Sensitivity for detection of Hodgkin lymphoma was similar for the two follow-up approaches. All of the relapses (40 of 40) were identified with FDG PET/CT (100%) and 39 of 40 relapses were identified with US/chest radiography (97.5%; P = .0001 for the equivalence test). US/chest radiography showed significantly higher specificity and positive predictive value than did PET/CT (96% [106 of 110] vs 86% [95 of 110], respectively; P = .02; and 91% [39 of 43] vs 73% [40 of 55], respectively; P = .01). Exposure to ionizing radiation was estimated to be 14.5 mSv for one PET/CT examination versus 0.1 mSv for one chest radiographic examination. Estimated cost per relapse diagnosed with routine PET/CT was 10-fold higher compared with that diagnosed with routine US/chest radiography., Conclusion: US and chest radiography are diagnostic tools that enable effective, safe, and low-cost routine surveillance imaging for patients at high risk of Hodgkin lymphoma relapse., (© RSNA, 2014.)

Isolated mediastinal amyloidosis is a rare condition. We report an unusual case of amyloid presented as an isolated mass, entirely confined within anterior mediastinum and FDG-avid, mimicking a neoplastic lesion. Because the differential diagnosis included several diseases as lymphoma, a biopsy via mediastinotomy was attended to avoid unnecessary sternotomy. The pathological results diagnosed to be an amyloidosis. The patient was asymptomatic and biopsy allowed an exact diagnosis, thus we decided against the complete excision. No monoclonal gammopathy and/or amyloid deposition were found. Thus, other treatments as high-dose melphalan and/or autologous stem cell transplantation were not indicated.

Primary smooth muscle tumors of the thyroid gland are extremely rare neoplasms. Due to their rarity, clinical case studies concerning management are lacking. According to a literature review, only 19 cases of primary thyroid leiomyosarcomas (TLs) have been reported. In the majority of patients, the prognosis is poor since adjuvant radiochemotherapy is ineffective on local recurrence and on long-term survival. In this study, we report the case of a 77-year-old male affected by a rapidly enlarging mass of the anterior neck, associated with bilateral lung metastases, and increasing dysphagia and dyspnea during the previous 6 months. A Tir4 neoplasm fine needle cytological diagnosis of the right thyroid lobe was reached and the patient underwent total thyroidectomy (TT). Definitive histological examination identified a TL. The patient succumbed 40 days later due to respiratory distress. A literature review was performed and TL differential diagnoses, management, including alternative treatment strategies, and adjuvant therapy were analyzed. TL is an aggressive rare mesenchymal malignant tumor. Although an improved multimodal approach is often necessary, TT and neck dissection represent the treatment of choice and are often the only possible therapy. Adjuvant radiochemotherapy appears to be ineffective and a high mortality rate is observed. TL remains a fatal tumor, and innovative and more effective therapeutic strategies to improve management and outcomes are required.

A case of acinic cell carcinoma of the breast is reported in a 26-year-old woman. She presented a lump in her right breast, that seemed to be a fibroadenoma. The open biopsy revealed a well-bordered fibroadenoma, together with a proliferation of cells characterized by serous acinar differentiation and eosinophilic cytoplasmic granules. Tumor cells stained for amylase, lysozyme, α-1-antichymotripsin, epithelial membrane antigen, S-100 protein, pan-cytokeratin, cytokeratin 7 and E-cadherin. Estrogen receptor, progesterone receptor, human epidermal growth factor receptor 2 overexpression, CD10, P63, smooth muscle actin, cytokeratin 5/6 were negative. The sentinel node was negative. 8 months after surgery she is in good clinical conditions without recurrence or metastases., (Copyright © 2014 Surgical Associates Ltd. Published by Elsevier Ltd. All rights reserved.)

Beckwith-Wiedemann syndrome (BWS) is an overgrowth disorder with increased risk of embryonal tumors, such as Wilms tumor, hepatoblastoma, neuroblastoma, and rhabdomyosarcoma. We report on a patient with BWS that developed a giant fibroadenoma of the breast that was surgically removed. The tumor relapsed 8 months after the surgery and the patient underwent partial mastectomy. Although the patient presented several clinical features of BWS, a molecular diagnosis was not achieved despite extensive molecular investigations on both blood and tumor tissue. A SNP array revealed a de novo 7p22.1 loss in both blood and breast tumor involving the mismatch repair gene PMS2 gene that may be potentially associated with the breast tumor. In conclusion, it remains unclear whether BWS patients have an increased risk of breast lesions or a yet unknown molecular defect is responsible for the rare occurrence of this tumor in BWS., (© 2013 Wiley Periodicals, Inc.)

Introduction: The breast and salivary gland tissue share embryologic and thus pathological similarities. Acinic cell carcinoma (ACC) is a typical tumor in salivary glands, but rarely arises in breast too. We reviewed 38 cases of mammary ACC reported in literature and our case, the first ACC born within a fibroadenoma., Materials and Methods: Data were collected by a research for the key words acinic cell carcinoma breast on Pubmed in March 2014, including a case treated in our department. All reviewed cases were compared for clinical approach and histological pattern., Results: To date 23 articles presenting cases of ACC of the breast are reported in literature. We included in our review 38 cases previously described and one new case. The histological pattern was predominantly solid with a microglandular structure. All the tumor cells were cytologically characterized by monotonous round cells with a finely granular, weakly eosinophilic, or clearly vacuolated cytoplasm. The most of the cells were intensely stained with anti-lysozime, anti-amylase, anti-α1-chimotripsin, anti-EMA and anti-S100 protein antisera. Immunohistochemistry was also performed to point out: estrogen receptor (ER), progesterone receptor (PR), androgen receptors (AR), human epidermal growth factor receptor 2 overexpression (HER2/neu), E-cadherin (E-cad), cytokeratin-7 (CK7), gross cystic disease fluid protein 15 (GCDFP15), smooth muscle actin (SMA)., Conclusion: ACC of the breast is a rare tumor, showing similarities with the salivary gland counterpart, above all in terms of good prognosis, and differences from the ordinary invasive breast carcinoma. Further investigations are needed to elucidate the true histogenesis and the correct treatment., (Copyright © 2014 Surgical Associates Ltd. Published by Elsevier Ltd. All rights reserved.)

Chordoma of thoracic vertebras is a very rare locally invasive neoplasm with low grade malignancy arising from embryonic notochordal remnants. Radical surgery remains the cornerstone of the treatment. We describe a case of multifocal T1-T2 chordoma, without bone and disc involvement, incidentally misdiagnosed as a paraganglioma, occurring in a 47-year-old male asymptomatic patient. Neoplasm was radically removed by an endocrine surgeon through a right extended cervicotomy. A preoperative reliable diagnosis of chordoma, as in the reported case, is often difficult. Radical surgery can provide a favorable outcome but, given the high rates of local recurrence of this neoplasm, a strict and careful follow-up is recommended. Although very rare, chordoma should be suggested in the differential diagnosis of the paravertebral cervical masses of unknown origin. Spine surgeon consultation and a FNB should be routinely included in the multidisciplinary preoperative work-up of these neoplasms.

Acinic cell carcinoma is a rare breast tumour belonging to salivary gland-like tumours of the breast. They are "triple-negative" breast cancers even if their biological behaviour seems to be more favourable. Herein we present an acinic cell carcinoma arising on a background of typical and atypical microglandular adenosis in a 58-year-old woman, along with a review of the literature.

Fine-needle aspiration (FNA) diagnosis of oncocytic lesions is challenging. In fact, oncocytic changes occur in inflammatory, hyperplastic, and neoplastic settings, including both benign and malignant tumors. The rare oncocytic variant of papillary thyroid carcinoma (PTC), shows papillae composed by cells with large oncocytic granular cytoplasm featuring clear PTC nuclear features. A morphological similar, but biologically distinct lesion, is the encapsulated papillary oncocytic neoplasia. Here, we first report on FNA, its cytological features together with histological, immunohistochemical, and molecular correlates., (Copyright © 2011 Wiley-Liss, Inc.)

We analyzed immunohistochemically the expression of CD24 and spliced variants of CD44v5 and v9 in invasive micropapillary carcinoma (IMPC) of the breast that is a rather aggressive tumor characterized by alteration of cells adhesion molecules, early lymph node metastases and poor prognosis. We analyzed 31 high-grade IMPCs and compared their expression to 22 high grade (G3) invasive ductal carcinomas of the breast (IDCs). We found a higher expression of CD24 in high-grade IMPCs with a peculiar inverted apical localization, compared to IDCs, showing a strong cytoplasmic staining; normal breast tissue resulted completely negative. IMPCs showed reduced expression of CD44v5 and CD44v9 compared with IDCs, but without a statistical significant difference. This study demonstrated that IMPC represents a distinct entity of breast carcinoma with high expression of CD24 with a typical inverted apical membrane pattern and reduction of CD44 isoforms v5 and v9, compared to IDCs. These features could explain the high lymph-vascular invasion propensity and higher metastatic capability of these tumors and could be a useful tool for a future targeted therapy., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

We recently published a study aiming to verify the frequency of amyloid deposits in the bone marrow of patients with multiple myeloma (MM) who did not present any signs or symptoms of systemic amyloidosis, applying the Congo red technique on bone marrow smears obtained by aspiration from the posterior iliac spine. The results suggested that nearly 40% of patients affected by MM may have amyloid deposits in their bone marrow. Subsequently, this finding has not been confirmed by another study performed with histological specimens of bone marrow in a similar clinical setting. To explain this discrepancy, we performed a comparative study on the bone marrows of 36 patients affected by MM, evaluated by both cytological and histological techniques. The results of this study confirm the high frequency of amyloid deposits in the bone marrow of patients affected by MM when the analysis is made on cytological smears, and indicate that the presence of amyloid on marrow smears is confirmed by core biopsies simultaneously performed in only 25% of cases. Should further studies confirm our findings, cytological assessment could be considered a sensitive technique to detect bone marrow amyloid deposits.

Background: Kidney transplantation is the therapy of choice in most cases of end stage renal disease. The purpose of the present study was to evaluate serum obestatin levels in kidney transplant recipients (Tx), compare levels in patients with renal failure (CKD) with those in healthy subjects (HS), and to assess the role of this hormone in energetic metabolism., Patients and Methods: A total of 95 subjects were studied: 40 were Tx; 35 had CKD and 20 were HS. Inclusion criteria were age>18years and good allograft function. Patients with an inflammatory disease or a diagnosis of cancer were excluded from the study., Results: Obestatin levels in Tx patients were significantly lower than in HS (3.5 [3-4.8] versus 11 [8.56-28.60] ng/mL; p<0.0001) and patients with CKD (3.5 [3-4.8] versus 4.7 [3, 5-6, 1] ng/mL; p=0.008). At univariate analysis, a direct correlation was found between obestatin and calcemia (p: 0.0001; r: 0.51), phosphoremia (p: 0.0005; r: 0, 46), calcium-phosphate product (p<0.0001; r:0.53), and parathormone (p: 0.01; r: 0.32), whereas significant inverse correlations were evidenced for BMI (p<0.0001; r: -0.52). At multivariate analysis, significance was maintained for the correlation between obestatin and phosphoremia (β=0.47; p=0.008), for the calcium-phosphate product (β=0.55; p=0.0005) and for BMI (β=-0.53; p=0.01)., Conclusion: Obestatin, present in lower levels in Tx patients than in CKD patients and HS, plays a role in energy metabolism, affecting BMI and the metabolism of calcium-phosphorus., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Because the lymphatic tissue of progressive transformation of germinal centers (PTGC) expresses CD20, rituximab treatment may prevent transformation to lymphoma of this rather atypical entity. We prospectively evaluated the efficacy of immunotherapy with rituximab (375 mg/m2 i.v. weekly for 4 consecutive weeks, followed by a single i.v. infusion of 375 mg/m2 every 3 months for 2 consecutive years) in 48 patients with biopsy-proven PTGC after Hodgkin lymphoma in complete remission post-induction therapy (4-6 courses of anthracycline-containing chemotherapy with radiotherapy). The event-free survival (EFS) of this series was compared with that of a historical cohort of 48 patients with PTGC developing after Hodgkin lymphoma in complete remission post-induction therapy, who underwent observation. At a median follow-up of 40 months, histology showed a malignancy in 27% of patients in the observation group (Hodgkin lymphoma, 13 patients) and in 2% of patients in the rituximab-protected group (non-Hodgkin lymphoma, one patient) (p ∼ 0.001). Rituximab was well tolerated in all treated patients. All relapses in the group not protected by immunotherapy involved the PTGC regions and non-contiguous nodal sites, which suggests that PTGC is a reservoir for malignant transformation and dissemination. The number needed to treat with rituximab to avoid one Hodgkin lymphoma relapse was four. Our study shows that prophylaxis with rituximab helps improve EFS in patients with PTGC and a history of Hodgkin lymphoma.

Objective: The aim of this study is to evaluate the correlation between the expression of some growth factors (GFs) and the tumor grade, recurrence and survival of brain glial and ependymal tumors., Material and Methods: The expression of vascular endothelial growth factor (VEGF), epidermal growth factor receptor (EGFR), tenascine, transforming growth factor (TGFbeta), isomeres, platelet-derived growth factor (PDGF) and p53 was studied in 40 primary brain tumors, both low-grade and high-grade, including astrocytomas, oligodendrogliomas, glioblastomas and ependymomas. The same GFs were also studied in 46 specimens of recurrent tumors from the same patients. The positivity and intensity of the immunohistochemical expression were correlated with the tumor grade, the interval and type of recurrence, and the survival., Results: The expression of all GFs, excepting TGFbeta1, TGFbetaRI and tenascine, was found to be correlated with the tumor grade in all tumors of both astroglial and oligodendroglial origin, whereas ependymomas showed significant differences only for EGFR. Low-grade (Grade II) tumors recurring as anaplastic (Grade III) forms showed GF expression rather similar to initially high-grade gliomas and significantly higher than that of low-grade (Grade II) tumors in both initial surgery and recurrence. Besides, low-grade (Grade II) tumors recurring as low-grade showed significantly longer median recurrence time (5.4 vs. 3.5 years) and better median survival (8.3 vs. 5.4 years) than those recurring as anaplastic forms (WHO III)., Conclusion: The immunohistochemical study of expression of VEGF, EGFR, TGFbeta2, TGFbeta3, PDGF and p53 in all low-grade (Grade II) brain gliomas at the first operation may help to differentiate cases with slower evolution and longer survival from those with higher potential of anaplastic transformation.

Aim: The use of a non-toxic tyrosine kinase receptor inhibitor, Imatinib Mesylate (IM), has become an ever-more common therapeutic alternative in some Kit (CD117) over-expressing neoplasms. As the treatment eligibility for these drugs hinges on CD117 expression, Kit immunostaining has recently been widely examined in various tumours. There are only limited data in the literature on the expression of c-kit expression in Wilms' Tumour. We examined CD117 expression in Wilms' tumour in order to correlate this marker with clinico-pathological data and to clarify its prognostic impact., Methods: This study included 40 cases of Wilms' tumour. Sections from paraffin-embedded tumour samples were immunostained by standard ABC technique using c-kit polyclonal antibody with antigen retrieval., Results and Conclusions: In the case of C-kit positive examples, the staining was focal, with patch distribution. On univariate analysis, significantly higher c-kit expression was observed in neoplasms in a more advanced stage of development than those in a less advanced stage (p=0.0055). In addition, over-expression of this marker was significantly correlated with the death of patients (p=0.0294) and recurrences of disease (p=0.0118). Moreover, all our Wilms' tumour anaplastic subtypes showed over-expression of c-kit and this was significantly higher than in favourable histology examples (p=0.0182). The results of multivariate analysis, instead, did not reveal any correlation of c-kit expression and prognosis. In our opinion these results could be due to the number of cases considered which is not particularly high. However, it seems likely that c-kit expression could be a secondary event related to tumour progression and could be influenced by chemotherapy and unfavourable histology.

This study reports a series of 19 extrapleural solitary fibrous tumors. The patients included 6 men and 13 women with age ranging from 27 to 86 years. Three patients showed local recurrence. In 2 tumors, a diagnosis of malignancy was made. All of the tumors were strongly positive for CD34, and 3 of them expressed high levels of progesterone receptor. Solitary fibrous tumors are fairly rare, occurring in many parts of the body, and their behavior is unpredictable.

Studies suggest Candida albicans infection has a negative effect on sperm function, including fertilizing ability. Assisted reproduction treatment using spermatozoa from a patient with unrecognized C. albicans infection did not result in fertilization. Preliminary evidence suggested an effect on sperm motility and apoptosis. This study was undertaken to evaluate the effects of experimentally induced C. albicans infection on motility, membrane mitochondrial potential (MMP), chromatin packaging and apoptosis [membrane phosphatidylserine (PS) externalization and DNA fragmentation] of spermatozoa isolated from normozoospermic healthy men. Motile spermatozoa were isolated by swim-up from 13 normal volunteers and exposed to increasing concentrations (0, 1000, 10,000, and 100,000 cfu/ml) of the fungus for 3 and 24 h. C. albicans was isolated from vaginal swabs, after identification, freshly prepared for experiments. Following incubation, sperm motility decreased significantly (P < 0.05 from 10,000 cfu/ml) and spermatozoa with reduced MMP or PS externalization, an early sign of apoptosis, increased in a time- and concentration-dependent manner. Sperm DNA fragmentation and chromatin integrity increased slightly after exposure to C. albicans, but the increase did not reach statistical significance. This study showed that C. albicans infection may decrease the functional competence of spermatozoa by reducing motility and MMP and by promoting molecular apoptosis mechanisms.

Background: Polymorphous hemangioendothelioma is a rare vascular tumor of borderline malignant potential and only 10 cases have been described in the literature so far., Methods and Results: We report a case of nodal and extranodal polymorphous hemangioendothelioma and review the literature. A 66-year-old man with an unremarkable past medical history was admitted for a left submandibular mass. The main nodule was composed of angiosarcoma-like areas mixed with angiomatous features; some vascular spaces with hobnail endothelium were seen. The tumor involved two adjacent lymph nodes. Immunohistochemically, the neoplastic cells were strongly positive for CD31 and vimentin. After a few months the tumor recurred and the patient was treated with radiation therapy., Conclusions: Polymorphous hemangioendothelioma is a distinct entity in the hemangioendothelioma group with its own clinical and histological features.