Israel Medical Association Journal

Quantitative chimerism testing has become an indispensable tool for following the course and success of allogeneic hematopoietic stem cell transplants. In this paper, we describe the current laboratory approach to quantitative chimerism testing based on an analysis of short tandem repeats, and explain why performing this analysis longitudinally is important and feasible. Longitudinal analysis focuses on relative changes appearing in the course of sequential samples, and as such exploits the ultimate potential of this intrinsically semi-quantitative platform. Such an analysis is more informative than single static values, less likely to be confused with platform artifacts, and is individualized to the particular patient. It is particularly useful with non-myeloablative conditioning, where mixed chimerism is common. When longitudinal chimerism analysis is performed on lineage-specific subpopulations, the sensitivity, specificity and mechanistic implications of the data are augmented. Importantly, longitudinal monitoring is a routinely feasible laboratory option because multiplex STR-PCR[1] kits are available commercially, and modern software can be used to perform computation, reliability testing, and longitudinal tracking in a rapid, easy to use format. The ChimerTrack© application, a shareware program developed in our laboratory for this purpose, produces a report that automatically summarizes and illustrates the quantitative temporal course of the patient’s chimeric status. Such a longitudinal perspective enhances the value of quantitative chimerism monitoring for decisions regarding immunomodulatory post-transplant therapy. This information also provides unique insights into the biological dynamics of engraftment underlying the fluctuations in the temporal course of a patient’s chimeric status.

Background: Decreased lacrimal gland output may cause dry eye syndrome. Using a rat model, we examined the feasibility of transplanting lacrimal gland cells from newborns.

Objectives: To restore lacrimal gland function in eyes with compromised tear production.

Methods: A model of dry eye in adult rats was developed by unilateral surgical removal of the main lacrimal gland. Tear secretion in both eyes was then assessed by masked Schirmer's test. Lacrimal gland tissue from newborn rats was transplanted into the fibrous connective tissue in which the lacrimal gland had been embedded. Masked Schirmer's test was repeated 4, 8 and 12 weeks after transplantation.

Results: Schirmer's test performed in 13 rats 10 days after unilateral lacrimal gland excision revealed significantly less wetting on the side with excised gland compared with the normal side (P < 0.003). The lack of secreting cells on the operated side was verified histologically. The reduction in tear secretion on the operated side remained significant for 8 weeks on average. In the six rats with transplanted lacrimal gland tissue however, there were no differences in tear reduction between the two eyes at 4, 8 or 12 weeks after the operation (P = 0.81, 0.56 and 0.8, respectively).

Conclusions: Transplantation of lacrimal gland tissue from newborn rats effectively restored eye wetting in this new model. Further research is needed to evaluate this new approach for treating lacrimal gland dysfunction. Using this model might also facilitate evaluation of potential clinical treatments for dry eyes.
 

Background: The model for end-stage liver disease is the best available predictor of waiting list mortality among liver transplant candidates.

Objectives: To validate the applicability of MELD[1] in Israel.

Methods: All candidates awaiting liver transplantation in our institution were followed prospectively since 2002. We measured the concordance (c-statistic) equivalent to the area under the receiver operating characteristic curve in order to assess the predictive power of MELD. Other independent mortality risk factors were identified by a separate multivariate analysis. Mortality rates within different MELD and Child‑Pugh‑Turcotte scores were compared to the original (United States) MELD data.

Results: Of 86 patients listed for transplantation, 40 were transplanted (36 in Israel and 4 abroad). Of the other 46 patients, 24 are alive and still listed, and 22 died (25%, ~7%/year). The area under the ROC[2] curve for MELD score was 0.79 (0.83 USA) compared to a CPT[3] score of 0.71 (O.76 USA). High MELD scores, occurrence of spontaneous bacterial peritonitis, and diagnosis of hepatocellular carcinoma were independent risk factors of mortality. Death rates per mid MELD score (20–29) were significantly higher than the USA results.

Conclusions: MELD is valid in Israel and superior to CPT in predicting waiting list mortality. Although longer waiting time due to organ scarcity is a key factor, death rates in the mid-range (10–29) MELD groups indicate further audit of the care of patients with end‑stage liver disease.

Objectives: To evaluate the national Israeli experience with lung transplantation in patients with CF[1].

Methods: We reviewed the medical charts of all CF patients who underwent lung transplantation between January 1996 and June 2005 at the two Israeli centers that performed this procedure.

Results: Eighteen transplantations were performed in 17 patients. Mean patient age at transplantation was 25.3 ± 9.1 years, and mean duration of follow-up in survivors (n=14) was 37.2 months (range 1–113 months). The actuarial survival rate was 88% at 1 year and 74% at 5 years. Pulmonary function, expressed as percent of predicted normal forced expiratory volume in 1 sec, improved from 22.4 ± 8.1% to 76 ± 16.8% at one year after transplantation. Bronchiolitis obliterans syndrome was diagnosed in 5 patients (29%), of whom 2 died and 2 are currently candidates for retransplantation. Median time to onset of BOS[2] was 34.2 months (range 17–64 months).

Conclusion: In Israel, the early and intermediate-term results of lung transplantation for cystic fibrosis are encouraging. BOS remains a major complication that threatens long-term outcome.

Objectives: To report 3 years national experience with orthotopic liver transplantation, and to evaluate patient and perioperative risk factors that could affect 1 year graft survival.

Methods: The study related to all 124 isolated adult liver transplantations performed in Israel between October 1997 and October 2000. Data were abstracted from the medical records. One-year graft survival was described using the Kaplan-Meier survival curve and three multivariate logistic regression models were performed: one with preoperative case-mix factors alone, and the other two with the addition of donor and operative factors respectively.

Results: Of the 124 liver transplantations performed, 32 failed (25.8%). The 1 year survival was lower than rates reported from both the United States and Europe, but the difference was not significant. Of the preoperative risk factors, recipient age ≥ 60 years, critical condition prior to surgery, high serum bilirubin and serum hemoglobin ≤ 10 g/dl were independently associated with graft failure, adjusting for all the other factors that entered the logistic regression equation. Extending the model to include donor and operative factors raised the C-statistic from 0.79 to 0.87. Donor age ≥ 40, cold ischemic time > 10 hours and a prolonged operation (> 10 hours) were the additional predictors for graft survival. A MELD score of over 18 was associated with a sixfold increased risk for graft failure (odds ratio = 6.5, P = 0.001).

Conclusions: Graft survival in Israel is slightly lower than that reported from the U.S. and Europe. Adding donor and operative factors to recipient characteristics significantly increased our understanding of 1 year survival of liver grafts.

The adult human heart has limited regenerative capacity and, therefore, functional restoration of the damaged heart presents a great challenge. Despite the progress achieved in the pharmacological and surgical treatment of degenerative myocardial diseases, they are still considered a major cause of morbidity and mortality in the western world. Repopulation of the damaged heart with cardiomyocytes represents a novel conceptual therapeutic paradigm but is hampered by the lack of sources for human cardiomyocytes. The recent derivation of pluripotent human embryonic stem cell lines may provide a solution for this cell sourcing problem. This review will focus on the derivation of the hESC[1] lines, their mechanism of self-renewal, and their differentiation to cardiomyocytes. The possible signals and cues involved in the commitment and early differentiation of cardiomyocytes in this model will be discussed as well as the molecular, structural and electrophysiologic characteristics of the generated hESC-derived cardiomyocytes. Finally, the hurdles and challenges toward fully harnessing the potential clinical applications of these unique cells will be described.

Objective: To present our algorithm of urgent and elective donor selection.

Methods: Urgent selection is expeditious and protocol‑based. Elective selection permits a comprehensive process. Both include medical, psychosocial and surgical-anatomic evaluations. Liver volumes and vascular anatomy are evaluated with computerized tomographic angiography. Informed consent is obtained after painstaking explanations. Independent institutional committees review and approve all cases.

Results: Between July 2003 and June 2004 we evaluated 43 potential live donors for 12 potential recipients (fulminant hepatic failure, n=5; chronic end-stage liver disease, n=6); primary graft non-function, n=1). Thirty-three candidates (76%) were excluded due to blood type incompatibility (n=14, 42%), incompatible anatomy (n=8, 24%) – including problematic volume distribution (n=2) or vascular anatomy (n=6) – psychosocial issues (n=4, 12%), or medical co-morbidity (n=7, 22%). Five recipients (FHF[1], n=4; chronic ESLD[2], n=1) were successfully transplanted from living donors. In the acute setting, two patients (FHF, PGNF[3]) died in the absence of an appropriate donor (cadaveric or living donor). In the elective group, one patient died of unexpected variceal bleeding and one received a cadaveric graft just before the planned living donor transplantation was performed. One candidate was transplanted overseas and two cases are scheduled. The ratio of compatibility for donation was 34% (10/29) for blood type-compatible candidates.

Conclusions: Donor selection for living donor liver transplantation is a complex, labor-intensive multidisciplinary process. Most exclusions are due to blood type incompatibility or anatomic details. Psychosocial aspects of these donations warrant special attention.

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[1] FHF = fulminant hepatic failure

[2] ESLD = chronic end-stage liver disease

[3] PGNF = primary graft non-function

Background: Pulmonary disease is the most frequent cause of morbidity and mortality in cystc fibrosis patients. New techniques such as non-invasive positive pressure ventilation have resulted in prolongation of life expectancy in CF[1] patients with end-stage lung disease.

Objectives: To determine the role of NIPPV[2] in CF patients awaiting lung transplantation.

Methods: Between 1996 and 2001 nine CF patients (5 females) with end-stage lung disease were treated with bi-level positive airway pressure ventilation in the "spontaneous" mode.

Results: The patients' mean age at initiation of BiPAP[3] was 15 years (range 13–40 years) and the mean duration of BiPAP usage was 8 months (range 3–16 months). Four patients underwent successful lung transplantation, three patients died while awaiting transplantation, and the remaining two are still on NIPPV while waiting for transplantation. Patients' body mass index increased significantly (P < 0.05) during BiPAP therapy (from 16.1 to 17.2 kg/m²). Blood pH, p_aCO₂, and bicarbonate improved significantly (from 7.31 to 7.38, 90.8 to 67.2 mmHg, and 48.9 to 40.3 mEq/L, respectively). Pulmonary function tests were not affected by BiPAP usage. The patients experienced a significant alleviation in morning headaches and improvement in quality of sleep (P < 0.003). There were no major complications during BiPAP usage.

Conclusions: We demonstrated that long-term NIPPV can stabilize and improve physiologic parameters such as ventilation, arterial blood gases and body mass index, as well as subjective symptoms such as sleep pattern, daily activity level, and morning headaches in CF patients with end-stage lung disease. Further prospectively controlled studies are needed to evaluate the potential of BiPAP therapy and its influence on morbidity and mortality in the post-lung transplantation period.

Background: The prognosis of patients with fulminant hepatic failure without timely liver transplantation is dismal. Given the limited availability of cadaveric organs for urgent transplantation in Israel, adult-to-adult living-donor segmental liver transplantation may be the only alternative.

Objectives: To report our initial experience with urgent lifesaving LDLT[1] in this unique scenario.

Methods: Three adult patients with FHF[2] (two of unknown etiology, one with paracetamol intoxication) were transferred from other institutions and admitted to our intensive care unit. Initial treatment and monitoring included intracranial pressure monitoring and hepatic dialysis using the Molecular Adsorbent Recirculating System. Expeditious potential donor selection included medical, psychosocial and surgical evaluation. Liver volume and vascular anatomic compatibility were assessed with computed tomography angiography.

Results: Between July and October 2003 we performed three procedures of urgent adult-to-adult LDLT. The donors (two uncles, one sister) underwent hepatic resection (two right lobes, one left lateral segment) and recovered well. The recipients underwent total hepatectomy with caval preservation, followed by lobar grafting. All recipients recovered and are alive with good liver function and without any neurologic complications.

Conclusions: Urgent adult-to-adult living-donor segmental liver transplantation can be performed safely and timely as a lifesaving procedure in the setting of comatose patients with FHF.

Since both major forms of diabetes involve inadequate function of pancreatic beta cells, intensive research is ongoing to better understand how beta cells perform their complex role of secreting the hormone insulin in response to physiologic needs. Identification and characterization of pancreatic transcription factors has revealed that they play a crucial role not only in maintenance of mature beta-cell function but also at multiple stages in pancreatic development. Furthermore, recent reports have revealed their potential to convert non-beta cells into insulin-producing cells, which in some cases can function to ameliorate diabetes in experimental animals. The ability to translate these successes to the clinic will require a detailed mechanistic understanding of the molecular basis of action of these proteins. Specific gene regulation in beta cells involves the action of multiple transcription factors recruited to the promoter and functioning synergistically to activate transcription, in part through recruitment of co-activator proteins and components of the basal transcriptional machinery. In addition, the process involves modification of chromatin structure, the details of which are beginning to be elucidated. Our ability to modulate gene expression patterns may lead to developing ways to provide an unlimited supply of functional beta cells for transplantation, permitting a dramatic improvement in therapeutic options for diabetes.