Francesco Nudo

Novel Stage Classification of Human Spermatogenesis Based on Acrosome Development

ABSTRACT To date, in the human seminiferous epithelium, only six associations of cell types have been distinguished, subdividing the epithelial cycle into six stages of very different duration. This hampers comparisons between studies on human and laboratory animals in which the cycle is usually subdivided into 12 stages. We now propose a new stage classification on basis of acrosomal development made visible by immunohistochemistry (IHC) for (pro)acrosin. IHC for acrosin gives results that are comparable to periodic acid Schiff staining. In the human too, we now distinguish 12 stages that differ from each other in duration by a factor of two at most. B spermatogonia are first apparent in stage I, preleptotene spermatocytes are formed in stage V, leptonema starts in stage VII, and spermiation takes place at the end of stage VI. A similar timing was previously observed in several monkeys. Stage identification by way of IHC for acrosin appeared possible for tissue fixed in formalin, Bouin fixative, diluted Bouin fixative, Cleland fluid, and modified Davidson fixative, indicating a wide applicability. In addition, it is also possible to distinguish the 12 stages in glutaraldehyde/osmium-tetroxide fixed/plastic embedded testis material without IHC for acrosin. The new stage classification will greatly facilitate research on human spermatogenesis and enable a much better comparison with results from work on experimental animals than hitherto possible. In addition, it will enable a highly focused approach to evaluate spermatogenic impairments, such as germ cell maturation arrests or defects, and to study details of germ cell differentiation.

De novo malignancies following liver transplantation: results from a multicentric study in central and southern Italy, 1990-2008.

OBJECTIVE The objective of this study was to quantify incidence rates (IR) and risks of de novo tumors (except nonmelanoma skin cancers) in patients who underwent orthotopic liver transplantation (OLT) in central and southern Italy. METHODS Data were collected on 1675 patients (75.5% males) who underwent OLT in six Italian transplantation centers in central and southern Italy (1990-2008). The time at risk of cancer (person years [PY]) was computed from OLT to the date of cancer diagnosis, death, or last follow-up, whichever occurred first. The number of observed cancer cases were compared with the expected one using data from population-based cancer registries. We computed gender- and age-standardized incidence ratios (SIRs) and 95% confidence intervals (CIs). RESULTS During 10,104.3 PYs (median follow-up, 5.2 years), 98 patients (5.9% of the total) were diagnosed with a de novo malignancy (for a total of 100 diagnoses). Twenty-two of these cancers were post-transplantation lymphoproliferative disorders (PTLD; 18 non-Hodgkin lymphoma [NHL] and 2 Hodgkins lymphoma [HL]), 6 were Kaposis sarcoma (KS), and 72 were solid tumors (19 head and neck [H&N], 13 lung, 11 colon-rectum, 6 bladder, and 4 melanoma). The overall incidence was 9.9 cases/10(3) PYs, with a 1.4-fold significantly increased SIR (95% CI, l.2-1.7). Significantly increased SIRs were observed for KS (37.3), PTLD (3.9), larynx (5.7), melanoma (3.1), tongue (7.1), and H&N (4.5) cancers. CONCLUSIONS These results confirmed that OLT patients are at greater risk for cancer, mainly malignancies either virus-associated or related to pre-existent factors (eg, alcohols). These observations point to the need to improve cancer surveillance after OLT. The on-going enrollment of patients in the present cohort study will help to elucidate the burden of cancer after OLT and better identify risk factors associated with its development.

Magnetic resonance-guided high-intensity focused ultrasound treatment of locally advanced pancreatic adenocarcinoma: preliminary experience for pain palliation and local tumor control.

PurposeThe purpose of this study was to evaluate the feasibility of magnetic resonance–guided focused ultrasound (MRgFUS) ablation for pain palliation and local tumor control in selected patients with unresectable primary pancreatic adenocarcinoma. Materials and MethodsAfter providing dedicated informed consent, 7 patients with histologically proven unresectable pancreatic adenocarcinoma underwent MRgFUS treatment on a dedicated 3-T unit featuring a dedicated ablation system. All lesions were evaluated for device accessibility before the treatment. Procedures of MRgFUS were performed with the patients under general anesthesia with constant controlled respiration. Clinical assessment included evaluation of symptom severity using a visual analog scale before and after the treatment. Imaging follow-up, including both computed tomographic and magnetic resonance examinations, was performed immediately after the treatment and at 3 and 6 months to evaluate the effects of MRgFUS on the targeted tumor and the occurrence, if any, of procedure-related complications. ResultsThe MRgFUS ablation was successfully performed in 6 patients; no adverse events were observed during or after the procedure. In a single patient, lesion accessibility was limited at treatment time, and the procedure was suspended. The visual analog scale score decreased in all patients from a mean (SD) of 7 (1) to 3 (1) after the treatment. Follow-up imaging results revealed negligible (n = 1) or no (n = 5) tumor regrowth within the ablation area. One patient died because of a metastatic disease 13 months after the treatment, whereas the other 5 are nonprogressing survivors at 6 and 8 months after the treatment. ConclusionsOur preliminary clinical experience suggests that MRgFUS is a feasible and repeatable ablative technique in selected patients with unresectable and device-accessible pancreatic adenocarcinoma.

Preharvest donor hyperoxia predicts good early graft function and longer graft survival after liver transplantation

A total of 44 donor/recipient perioperative and intraoperative variables were prospectively analyzed in 89 deceased‐donor liver transplantations classified as initial good graft function (IGGF) or initial poor graft function (IPGF) according to a scoring system based on values obtained during the 1st 72 postoperative hours from the serum alanine aminotransferase (ALT) concentration, bile output, and prothrombin activity. The IGGF compared with the IPGF group showed: 1) longer graft (P = .002) and patient (P = .0004) survival; 2) at univariate analysis, a higher (mean [95% confidence interval]) preharvest donor arterial partial pressure of oxygen (PaO2) (152 [136‐168] and 104 [91‐118] mmHg, respectively; P = .0008) and arterial hemoglobin oxygen saturation (97.9 [97.2‐98.7] and 96.7 [95.4‐98.0]%, respectively; P = .0096), a lower percentage of donors older than 65 years (13 and 33%, respectively; P = .024), a lower percentage of donors treated with noradrenaline (16 and 41%, respectively; P = .012). At multivariate analysis, IGGF was associated positively with donor PaO2 and negatively with donor age greater than 65 years and with donor treatment with noradrenaline. Independently from the grouping according to initial graft function, graft survival was longer when donor PaO2 was >150 mmHg than when donor PaO2 was ≤150 mmHg (P = .045). In conclusion, preharvest donor hyperoxia predicts IGGF and longer graft survival. (Liver Transpl 2005;11:140–151.)

Preoperative donor scores and postoperative early measures of graft function: relevance to the outcome of liver transplantation.

BACKGROUND Several donor and recipient parameters play a role in the determination of post-liver transplant allograft function. The identification of prognostic indices presents great implications for correct allocation of donors and more targeted recipient management. The aim of our review was to detect the role of preoperative scoring systems and early postoperative measures of graft function as predictive factors for the development of graft failure and recipient death. METHODS We stratified a cohort of 97 patients in two groups according to a 1-year functional (Group A; n = 72) versus non-functional (Group B; n = 25) status of the allograft. RESULTS Patients in group B showed higher preoperative Model for End-stage Liver Disease (MELD) values, longer warm ischemia times, reduced bile outputs and increased peak values of transaminases and INR content within the first 3 days after transplantation. Group B showed 48% of patients with initial poor graft function. The parameters which resulted in a significant prediction of graft loss by multivariate analysis were MELD (P = .012); postoperative day 1 serum alanine aminotransferase (ALT) (P < .0001) and day 3 ALT (P = .003). The predictive factors for patient death were postoperative day 1 serum ALT (P < .0001) and day 3 ALT (P = .001). CONCLUSIONS MELD score was a useful preoperative parameter for the prediction of post-transplant graft survival. Early ALT values predicted both graft and recipient survivals. Minimization of parameters related to their peaks (warm ischemia time) may improve graft and patients survival rates.

Spermatogonial kinetics in humans

The human spermatogonial compartment is essential for daily production of millions of sperm. Despite this crucial role, the molecular signature, kinetic behavior and regulation of human spermatogonia are poorly understood. Using human testis biopsies with normal spermatogenesis and by studying marker protein expression, we have identified for the first time different subpopulations of spermatogonia. MAGE-A4 marks all spermatogonia, KIT marks all B spermatogonia and UCLH1 all Apale-dark (Ap-d) spermatogonia. We suggest that at the start of the spermatogenic lineage there are Ap-d spermatogonia that are GFRA1High, likely including the spermatogonial stem cells. Next, UTF1 becomes expressed, cells become quiescent and GFRA1 expression decreases. Finally, GFRA1 expression is lost and subsequently cells differentiate into B spermatogonia, losing UTF1 and acquiring KIT expression. Strikingly, most human Ap-d spermatogonia are out of the cell cycle and even differentiating type B spermatogonial proliferation is restricted. A novel scheme for human spermatogonial development is proposed that will facilitate further research in this field, the understanding of cases of infertility and the development of methods to increase sperm output. Summary: Marker protein expression pattern analysis improves understanding of human spermatogonial development and stem cell renewal, facilitating further research and providing insight into fertility problems.

Does Caval Reconstruction Technique Affect Early Graft Function after Liver Transplantation? A Preliminary Analysis

BACKGROUND In the past decades, the inferior vena cava (IVC) reconstruction technique has undergone several evolutions, such as biopump, piggyback technique (PB), and laterolateral approach (LLPB). Several advantages are reported comparing the PB technique to biopump use. However, comparison between PB and LLPB has not been as well investigated. The aim of this study was to compare the results in terms of immediate graft function and intermediate graft survival among 3 subgroups characterized by distinct caval reconstruction techniques. METHODS We retrospectively analyzed a cohort of 200 consecutive adult patients who underwent liver transplantation from January 2001 to December 2009. The patients were stratified according to 3 caval reconstructive techniques: biopump (n=135), PB (n=32) and LLPB (n=33). RESULTS The LLPB group showed the shortest cold and warm ischemia times and the best immediate postoperative graft function. Survival analysis revealed LLPB patients to present the best 1-year graft survival rates: namely, 90.9% versus 75.0% and 74.1% among the PB and biopump groups, respectively (log-rank tests: LLPB vs biopump: P=.03; LLPB vs PB: P=.05). In our experience, LLPB showed the best graft survivals with an evident reduction in both cold and warm ischemia times. However, it is hard to obtain an irrefutable conclusion owing to the retrospective nature of this study, the small sample, and the different periods in which the groups were transplanted. CONCLUSIONS LLPB technique was a safe procedure that minimized the sequelal of ischemia-reperfusion damage. This technique yielded results superior to venovenous bypass. No definitive conclusions can to be obtained in this study comparing classic PB or LLPB.

Simultaneous pancreas-kidney transplantation: a single-center experience and prospective analysis.

In patients with end-stage chronic kidney disease (CKD) and type 1 diabetes mellitus (DM 1), simultaneous pancreas-kidney (SPK) transplantation is currently considered the gold standard therapy. The aim of this study was to analyze and report the long-term clinical outcomes of the 23 SPK transplantations performed at our institution over an 84-month period (January 1, 2000 to December 31, 2006). A prospective analysis of these patients included donor, recipient, and transplantation characteristics. The only requirements for transplantation were blood group compatibility and a negative cross-match. Bladder drainage via pancreaticoduodenocystostomy was performed in all of the patients. Due to a pulmonary embolus 1 patient (4.3%) died at 2 months. The actuarial patient survival rates at 3 months and 1, 3, and 5 years were 95.6%. Causes for the renal graft loss were chronic allograft nephropathy in 3 cases (13%) and death of the patient in 1 case (4.3%). The actuarial censored renal allograft survival rates at 3 months and at 1 year were 100%, and at 3 and 5 years were 91.3%. Causes for the renal graft loss were chronic rejection in 1 case (4.3%) and patient death in 1 case (4.3%). The actuarial censored pancreatic allograft survival rates at 3 months and at 1 and 3 years were 100%, and at 5 years was 95.6%. The results of this work add further evidence that SPK is the gold standard therapy for selected patients with end-stage CKD due to DM 1.

Head and neck and esophageal cancers after liver transplant: results from a multicenter cohort study. Italy, 1997–2010

This study quantified the risk of head and neck (HN) and esophageal cancers in 2770 Italian liver transplant (LT) recipients. A total of 186 post‐transplant cancers were diagnosed—including 32 cases of HN cancers and nine cases of esophageal carcinoma. The 10‐year cumulative risk for HN and esophageal carcinoma was 2.59%. Overall, HN cancers were nearly fivefold more frequent in LT recipients than expected (standardized incidence ratios ‐ SIR=4.7, 95% CI: 3.2–6.6), while esophageal carcinoma was ninefold more frequent (SIR=9.1, 95% CI: 4.1–17.2). SIRs ranged from 11.8 in LT with alcoholic liver disease (ALD) to 1.8 for LT without ALD for HN cancers, and from 23.7 to 2.9, respectively, for esophageal carcinoma. Particularly elevated SIRs in LT with ALD were noted for carcinomas of tongue (23.0) or larynx (13.7). Our findings confirmed and quantified the large cancer excess risk in LT recipients with ALD. The risk magnitude and the prevalence of ALD herein documented stress the need of timely and specifically organized programs for the early diagnosis of cancer among LT recipients, particularly for high‐risk recipients like those with ALD.

Inferior vena cava interruption in a liver transplantation deceased donor

Inferior vena cava (IVC) abnormalities occur in lessthan 1% of patients, being their incidence higher incase of congenital heart diseases (1). IVC embryologicdevelopment is a complex process involving the for-mation and regression of different venous systems (2).Pre-existing IVC abnormalities have been welldescribed in liver transplant recipients (3) but not inthe case of donors. We report the case of a 62-year-old Caucasian man who was considered eligible formultiorgan donation after diagnosis of cerebral death.During the organ procurement, the surgical teamnoted a retrohepatic IVC interruption with azygos con-tinuation (Fig. 1); this anomaly was further confirmedat the back-table surgery, in which only the suprahe-patic part of the IVC was present (Fig. 2). After surgi-cal evaluation, the graft was retained suitable for livertransplantation. The recipient was a 64-year-old manwith a cryptogenic cirrhosis. IVC reconstruction wasdone preserving the recipient IVC and performing alatero-lateral anastomosis with stapler closure of thedonor sovrahepatic IVC. Total surgical time was360 min, with cold and warm ischaemia times of 385and 51 min respectively. After a 3-year follow-up, therecipient is alive with no vascular complication. The la-tero-lateral anastomosis performed in this case is dis-played in Figure 3. IVC abnormalities are uncommonlydiscovered during organ procurement: however, theirpresence does not affect the possibility to perform aliver transplantation.