EpiMethylTag: simultaneous detection of ATAC-seq or ChIP-seq signals with DNA methylation.

Activation of regulatory elements is thought to be inversely correlated with DNA methylation levels. However, it is difficult to determine whether DNA methylation is compatible with chromatin accessibility or transcription factor (TF) binding if assays are performed separately. We developed a fast, low-input, low sequencing depth method, EpiMethylTag, that combines ATAC-seq or ChIP-seq (M-ATAC or M-ChIP) with bisulfite conversion, to simultaneously examine accessibility/TF binding and methylation on the same DNA. Here we demonstrate that EpiMethylTag can be used to study the functional interplay between chromatin accessibility and TF binding (CTCF and KLF4) at methylated sites.

Discovery of a periosteal stem cell mediating intramembranous bone formation.

Bone consists of separate inner endosteal and outer periosteal compartments, each with distinct contributions to bone physiology and each maintaining separate pools of cells owing to physical separation by the bone cortex. The skeletal stem cell that gives rise to endosteal osteoblasts has been extensively studied; however, the identity of periosteal stem cells remains unclear1-5. Here we identify a periosteal stem cell (PSC) that is present in the long bones and calvarium of mice, displays clonal multipotency and self-renewal, and sits at the apex of a differentiation hierarchy. Single-cell and bulk transcriptional profiling show that PSCs display transcriptional signatures that are distinct from those of other skeletal stem cells and mature mesenchymal cells. Whereas other skeletal stem cells form bone via an initial cartilage template using the endochondral pathway4, PSCs form bone via a direct intramembranous route, providing a cellular basis for the divergence between intramembranous versus endochondral developmental pathways. However, there is plasticity in this division, as PSCs acquire endochondral bone formation capacity in response to injury. Genetic blockade of the ability of PSCs to give rise to bone-forming osteoblasts results in selective impairments in cortical bone architecture and defects in fracture healing. A cell analogous to mouse PSCs is present in the human periosteum, raising the possibility that PSCs are attractive targets for drug and cellular therapy for skeletal disorders. The identification of PSCs provides evidence that bone contains multiple pools of stem cells, each with distinct physiologic functions.

DeTiN: overcoming tumor-in-normal contamination.

Comparison of sequencing data from a tumor sample with data from a matched germline control is a key step for accurate detection of somatic mutations. Detection sensitivity for somatic variants is greatly reduced when the matched normal sample is contaminated with tumor cells. To overcome this limitation, we developed deTiN, a method that estimates the tumor-in-normal (TiN) contamination level and, in cases affected by contamination, improves sensitivity by reclassifying initially discarded variants as somatic.

Chromosomal Instability as a Driver of Tumor Heterogeneity and Evolution.

Large-scale, massively parallel sequencing of human cancer samples has revealed tremendous genetic heterogeneity within individual tumors. Indeed, tumors are composed of an admixture of diverse subpopulations-subclones-that vary in space and time. Here, we discuss a principal driver of clonal diversification in cancer known as chromosomal instability (CIN), which complements other modes of genetic diversification creating the multilayered genomic instability often seen in human cancer. Cancer cells have evolved to fine-tune chromosome missegregation rates to balance the acquisition of heterogeneity while preserving favorable genotypes, a dependence that can be exploited for a therapeutic benefit. We discuss how whole-genome doubling events accelerate clonal evolution in a subset of tumors by providing a viable path toward favorable near-triploid karyotypes and present evidence for CIN-induced clonal speciation that can overcome the dependence on truncal initiating events.

Mutational heterogeneity in cancer and the search for new cancer-associated genes.

Major international projects are underway that are aimed at creating a comprehensive catalogue of all the genes responsible for the initiation and progression of cancer. These studies involve the sequencing of matched tumour-normal samples followed by mathematical analysis to identify those genes in which mutations occur more frequently than expected by random chance. Here we describe a fundamental problem with cancer genome studies: as the sample size increases, the list of putatively significant genes produced by current analytical methods burgeons into the hundreds. The list includes many implausible genes (such as those encoding olfactory receptors and the muscle protein titin), suggesting extensive false-positive findings that overshadow true driver events. We show that this problem stems largely from mutational heterogeneity and provide a novel analytical methodology, MutSigCV, for resolving the problem. We apply MutSigCV to exome sequences from 3,083 tumour-normal pairs and discover extraordinary variation in mutation frequency and spectrum within cancer types, which sheds light on mutational processes and disease aetiology, and in mutation frequency across the genome, which is strongly correlated with DNA replication timing and also with transcriptional activity. By incorporating mutational heterogeneity into the analyses, MutSigCV is able to eliminate most of the apparent artefactual findings and enable the identification of genes truly associated with cancer.

Depletion of T regulatory cells through selection of CD127-positive cells results in a population enriched in memory T cells: implications for anti-tumor cell therapy.

BACKGROUND: Donor lymphocyte infusions can induce remissions in patients with relapse after allogeneic hematopoietic stem cell transplantation. Nevertheless, some grafted patients never display any signs of alloreactivity, either following allogeneic hematopoietic stem cell transplantation or after donor lymphocyte infusions. Consequently, they do not develop graft-versus-host disease and frequently do not respond to donor lymphocyte infusions. In a recently published clinical trial, we observed that elimination of CD4(+)CD25(+)Foxp3(+) natural regulatory T cells from the donor lymphocyte product could improve alloreactivity and the associated anti-tumor effect in a small proportion of patients with relapsed hematologic malignancies. Here, we aimed to improve the effect of donor lymphocyte infusion by modifying the procedure for depletion of T regulatory cells. DESIGN AND METHODS: We directly compared depletion of regulatory T cells from human peripheral blood mononuclear cells achieved by selection of CD127-positive cells or by selection of CD25-negative cells. We tested the manipulated products (i) in vitro in mixed lymphocyte reactions and against pathogen-derived recall antigens and (ii) in vivo in experimental graft-versus-host disease. RESULTS: In vitro, we found that depletion of regulatory T cells through CD127 positive selection improved both alloreactive and pathogen-specific immune responses. In vivo, we observed accelerated donor T-cell division and enhanced graft-versus-host disease due to efficient regulatory T-cell depletion accompanied by enrichment in memory T cells. CONCLUSIONS: Our results show that the strategy of CD127 positive selection is an efficient way of eliminating regulatory T cells from donor lymphocyte infusions and improves alloreactivity. This supports the investigation of CD127 positive selection in place of elimination of CD25-positive cells for clinical applications.

MicroRNAs in mutagenesis, genomic instability, and DNA repair.

MicroRNAs (miRNAs) are aiding our understanding of cancer biology, and are now coming close to therapeutic use as well. Here, we focus specifically on the interaction between miRNAs and genomic instability. MiRNA regulation is essential to many cellular processes, and escape from this regulatory network seems to be a common characteristic of malignant transformation. Genomic instability may preferentially target miRNAs either because of selective pressure or because of inherent vulnerability related to their location near fragile sites. Furthermore, disruption of miRNA processing elements affords a more global release from miRNA regulation. Finally, we review how miRNAs function as both effectors and modulators of the DNA damage response, intricately weaved with traditional elements such as ATM, P53, and MMR. Thus, miRNAs are important substrates for genomic instability and play a crucial role in cellular DNA sensing and repair mechanisms.

Immune reconstitution is preserved in hematopoietic stem cell transplantation coadministered with regulatory T cells for GVHD prevention.

Recipient-specific regulatory T cells (rsTreg) can prevent graft-versus-host disease (GVHD) by inhibiting donor T-cell expansion after hematopoietic stem cell transplantation (HSCT) in mice. Importantly, in adult humans, because of thymus involution, immune reconstitution during the first months after HSCT relies on the peripheral expansion of donor T cells initially present in the graft. Therefore, we developed a mouse model of HSCT that excludes thymic output to study the effect of rsTreg on immune reconstitution derived from postthymic mature T cells present within the graft. We showed that GVHD prevention with rsTreg was associated with improvement of the limited immune reconstitution compared with GVHD mice in terms of cell numbers, activation phenotype, and cytokine production. We further demonstrated a preserved in vivo immune function using vaccinia infection and third-party skin-graft rejection models, suggesting that rsTreg immunosuppression was relatively specific of GVHD. Finally, we showed that rsTreg extensively proliferated during the first 2 weeks and then declined. In turn, donor Treg proliferated from day 15 on. Taken together, these results suggest that rsTreg GVHD prevention is associated with improved early immune reconstitution in a model that more closely approximates the biology of allogeneic HSCT in human adults.

Regulatory T cell content in the bone marrow graft does not predict the occurrence of acute GVHD.

The subpopulation of regulatory T cells (Treg) was shown to play a key role in alloreactive responses. In allogeneic hematopoietic stem cell transplantation, several groups tested whether Treg content in transplants correlates with graft-versus-host disease (GVHD) with controversial results. In a retrospective study of 49 consecutive HLA-matched sibling transplantations, we studied the relationship between Treg content in bone marrow transplants and acute GVHD (aGVHD) occurrence. We observed a large variability in Treg in bone marrow grafts. However, contrary to previous observations in peripheral blood stem cells transplantation, we report that the Treg content of allogeneic bone marrow transplantation did not predict the occurrence of aGVHD.

Causes and predictive factors of mortality in a cohort of patients with hepatitis C virus-related cryoglobulinemic vasculitis treated with antiviral therapy.

OBJECTIVE: Hepatitis C virus (HCV)-associated mixed cryoglobulinemia (MC) vasculitis is an autoimmune disorder with significant morbidity and mortality. Renal involvement was associated with an increased mortality, and was the most common cause of death; these data were obtained before effective antiviral treatment was available. We studied causes of death and predictive factors in patients with HCV-associated MC vasculitis treated with antivirals. METHODS: Case histories of 85 patients with HCV-associated MC vasculitis treated in a single center between 1990 and 2006 were retrospectively reviewed. Prognostic factors affecting mortality were studied by comparing 23 patients who died with 62 survivors, using the Cox model regression analysis. RESULTS: The most common cause of death was infection, accounting for 34.7%, followed by endstage liver disease in 30.4% (including 4 patients with hepatocellular carcinoma), and cardiovascular disease in 17.4% of patients. Endstage renal disease accounted for only 8.7% of deaths, as did central nervous system vasculitis and nonhepatic malignancy. Increased mortality was strongly associated with immunosuppressive treatment [hazard ratio (HR) 6.51, 95% CI 2.75-15.37], cutaneous ulcers (HR 5.37, 95% CI 1.79-16.14), and renal insufficiency (HR 3.25, 95% CI 1.37-7.72). A 2 log10 decrease in HCV viral load at month 3 after starting antiviral treatment was associated with decreased mortality (HR 0.39, 95% CI 0.16-0.95). CONCLUSION: While renal involvement is still associated with poorer prognosis, infectious processes are now the most common cause of death in HCV cryoglobulinemia vasculitis. Immunosuppressive treatment is associated with an increased risk of death, independently from disease severity. Response to antiviral treatment is associated with significantly reduced mortality risk.

Role of air-medical evacuation in mass-casualty incidents--a train collision experience.

BACKGROUND: On 21 June 2005, a passenger train collided with a truck near Revadim, Israel. The collision resulted in a multiple-scene mass-casualty incident in an area characterized by difficult access and a relatively long distance from trauma centers. A major disaster response was initiated by civilian and military medical forces including the Israeli Air Force (IAF) Search and Rescue teams. The air-medical evacuation from the accident site to the trauma centers, the activities of the airborne medical teams, and the lessons learned from this event are described in this report. METHODS: A retrospective analysis of data gathered from relevant elements that participated in management, treatment, and evacuation from the accident site was conducted. RESULTS: The accident resulted in 289 injured passengers and seven of the injured were killed. Six helicopters (performing nine sorties) participated. Helicopters evacuated trauma victims and aided in transporting air-medical teams to the site of the collision. Overall, 35 trauma victims (10 urgent) were evacuated by air to trauma centers. The length of time between the first helicopter landing and completion of the air evacuation was 83 minutes. The air-medical evacuation operation was controlled by the commander of the IAF Search and Rescue. Different crew compositions were set in real time. CONCLUSIONS: Air-medical evacuation during this unique event enabled prompt transportation of casualties from the scene to trauma centers and provided reasonable distribution of patients between various centers in the region. This operation highlighted the necessity for flexibility in medical decision-making and the need for nonconventional solutions regarding crew compositions during management of an airborne evacuation in similar settings. Air-medical evacuation should be considered as a part of responses to mass-casualty incidents, especially when the site is remote or characterized by accessibility difficulties.

Correlation of clinical and virologic responses to antiviral treatment and regulatory T cell evolution in patients with hepatitis C virus-induced mixed cryoglobulinemia vasculitis.

OBJECTIVE: Mixed cryoglobulinemia (MC) vasculitis is an autoimmune disorder associated with chronic hepatitis C virus (HCV) infection. We previously reported that MC vasculitis is associated with a quantitative defect of peripheral blood regulatory T cells. The aim of this study was to prospectively evaluate the evolution of this defect during the course of antiviral treatment. METHODS: Treg cell frequencies and numbers were analyzed in 131 patients with chronic HCV infection (including 66 with MC vasculitis) and 20 healthy volunteer donors. Measurements were taken before, during, and after treatment with PEGylated interferon alfa-2b plus ribavirin. RESULTS: At baseline, patients with MC vasculitis had a significantly lower frequency and number of Treg cells than did patients without MC vasculitis. Complete remission of MC vasculitis following antiviral treatment was associated with a significant increase in Treg cell levels compared with baseline. In contrast, Treg cell levels in nonresponders or partial responders, which did not differ from those in complete responders at baseline, remained unchanged over the course of the study. CONCLUSION: The strong positive correlation between clinical responses and Treg cell levels provides further support for the central role of Treg cells in the pathogenesis of HCV-induced MC vasculitis and emphasizes the dual role of Treg cells in chronic HCV infection: while Treg cells may hinder viral elimination, they also limit autoimmune injury.

The prevalence of gastrointestinal diseases in Israeli adolescents and its association with body mass index, gender, and Jewish ethnicity.

OBJECTIVE: The objectives of this study were to describe gastrointestinal (GI) disease prevalence in Israeli adolescents, and possible associations between prevalence and body mass index (BMI), sex and Jewish ethnicity. DESIGN: A retrospective analysis of screening for GI disease performed by the Israeli Defense Forces recruiting office between 1998 and 2003. SETTING: Screening was performed at the recruiting office, and included detailed history and physical examination performed by a general practitioner. Further testing was performed as needed and the final diagnosis was established by a gastroenterologist at the recruiting office. PATIENTS: Seventeen-year-old Israeli nationals. RESULTS: During the study period, 466,855 (58.5% male) adolescents were screened for GI disease. Peptic ulcer disease, irritable bowel syndrome, and nonulcer dyspepsia were the most prevalent disorders affecting 466/10, 460/10, and 296/10, respectively. There was an increase in the prevalence of lactose intolerance during the study period and also an increase in the prevalence of peptic ulcer disease in females. The prevalence of inflammatory bowel disease has also increased from 100/10 to 149/10, although this trend failed to reach statistical significance (P=0.097). Higher BMI was associated with statistically significant higher prevalence rates of gastroesophageal reflux disease (P<0.05). A stronger association in females was found in gallbladder disease (P<0.001). Lower BMI was associated with higher prevalence rates of irritable bowel syndrome (P<0.001), and higher rates of inflammatory bowel disease and lactose intolerance in males (P<0.01 and <0.001, respectively). CONCLUSIONS: GI diseases are not uncommon among adolescents, and for some disorders prevalence is rising. The association between BMI and prevalence has been further clarified.

Detection of occult lung impairment in welders by induced sputum particles and breath oxidation.

BACKGROUND: We evaluated particulate matter in combined induced sputum (IS) and oxidation in exhaled breath condensate (EBC) to test whether underlying inflammatory changes are present in asymptomatic welders. METHODS: Thirty welders from the Israel Defense Forces exposed to aluminum/iron (Group 1) or to cadmium/chromium/iron/lead/nickel (Group 2, N = 16) and 27 non-exposed administrators were studied. IS was recovered, particle size distribution, hydrogen peroxide and pH were measured, and exhaled breath condensate was collected. RESULTS: Group 2 had a higher % neutrophils than all other participants (P = 0.0001) and a higher % particles >2 microm in diameter (P = 0.0017). Percent particles and years of exposure highly correlated (P = 0.051). All welders EBC samples had higher concentrations of hydrogen peroxide than controls (P = 0.0001). pH was lower only for Group 2 (P = 0.0001). CONCLUSIONS: Combined IS and EBC measurements detect underlying inflammation in airways of asymptomatic welders. It emerged that airway inflammation is present in asymptomatic welders, and that the particle burden, inflammatory cells, and level of oxidative stress are a function of the type and the duration of welding. Am. J. Ind. Med. 51:503-511, 2008.

Relapse of hepatitis C virus-associated mixed cryoglobulinemia vasculitis in patients with sustained viral response.

OBJECTIVE: To investigate the clinical characteristics, outcomes, and results of hepatitis C virus (HCV) RNA analyses in a group of patients with HCV-associated mixed cryoglobulinemia (MC) vasculitis who experienced a relapse of vasculitis despite achieving a sustained viral response to treatment with antiviral agents. METHODS: HCV RNA testing was performed by the transcription-mediated amplification (TMA) method in sera and cryoprecipitates (detection limit 2.5 IU/ml). HCV replication was assessed in peripheral blood mononuclear cells (PBMCs) by a modified real-time polymerase chain reaction assay (detection limit 15 IU/10(6) cells). RESULTS: We identified 8 patients with relapse of HCV-MC vasculitis despite their having achieved a sustained viral response to treatment. Relapse appeared early after the end of treatment (mean +/- SD 2.5 +/- 3.5 months) and included mainly purpura (n = 7) and arthralgia (n = 5). Relapse was associated with an increase in serum cryoglobulin levels as compared with end-of-treatment levels (mean +/- SD 0.3 +/- 0.09 gm/liter and 0.08 +/- 0.04 gm/liter, respectively; P < 0.01) and a decrease in C4 levels. In most patients, the relapse was brief, and the MC vasculitis manifestations subsided. A search for HCV RNA by TMA was negative in all patients tested (7 of 8 patients), both in sera and in cryoprecipitates. HCV replication was not found in PBMCs from any of the patients tested (6 of 8 patients). In 3 patients, the MC vasculitis symptoms persisted and were associated with elevated cryoglobulin levels. B cell lymphoma was diagnosed in 2 of these 3 patients. CONCLUSION: Relapse of MC vasculitis does occur in a few patients with HCV infection, despite achieving a sustained viral response, and this relapse is not related to persistence of virus. Relapse is short-lived and may be induced by the withdrawal of interferon alfa therapy. However, in patients with persistent MC vasculitis symptoms, a different underlying condition should be considered, especially B cell lymphoma.

Physical examination and ECG screening in relation to echocardiography findings in young healthy adults.

BACKGROUND AND AIMS: Cardiovascular screening in young adults is an important tool in many occupational settings. Our aim was to test whether screening physical examination and ECG influence the rate of abnormal echocardiogarphic findings in young healthy subjects. METHODS: Consecutive echocardiography results of 18- to 20-year-old flight candidates were analyzed retrospectively. Echocardiographies were performed as part of a screening protocol, which includes ECG, physical examination and referral for echocardiography for any positive finding. A second stage includes universal echocardiography for all candidates. RESULTS: 1,066 subjects were evaluated; 489 subjects underwent echocardiography following referral because of abnormal auscultatory or ECG findings. Findings (mostly mild valvular insufficiencies) were demonstrated in 12.7%, with only 0.6% of subjects disqualified. In subjects who underwent universal echocardiography (n = 577), findings (mostly mild valvular insufficiencies) were detected in 18%, with only 0.5% of subjects disqualified. CONCLUSIONS: The rate of significant echocardiography findings is extremely low in this young and healthy population. The presence of abnormal findings on either physical examination or ECG screening was not demonstrated to alter the rate of abnormal echocardiographic findings. We suggest that the low yield of screening should be weighed against the cost of an unidentified congenital cardiac lesion in the specific setting.

Celiac disease diagnosed in the elderly.

BACKGROUND AND AIMS: In the past 20 years, a growing proportion of new cases of celiac disease (CD) are diagnosed in adults and in patients with extraintestinal manifestations. Our understanding of the extremely wide spectra of manifestations and the profound effects on elderly patients is improving. Nevertheless, CD is still underdiagnosed in elderly patients. In this study, we describe a case series of CD patients diagnosed after the age of 60. METHODS: A retrospective chart review was preformed in cases of CD diagnosed after the age of 60. Patients were included if they had positive serology and histologic findings compatible with CD. Eligible patients were reinterviewed, and demographic, clinical, and laboratory information were recorded. RESULTS: During the study period, 7 patients with CD diagnosed after the age of 60 were identified. The most common presenting findings were weight loss, iron deficiency anemia, and diarrhea. Two patients suffered from severe early osteoperosis and 2 additional patients had elevated liver function tests. Neurologic manifestation was suspected in 3 cases. Two female patients presented with cognitive decline that was attributed to Alzheimer dementia but ameliorated after the initiation of gluten-free diet. The third patient had peripheral neuropathy that completely resolved after the initiation of gluten-free diet. Median lag in diagnosis was 8 years. Diet treatment led to complete resolution of symptoms in most cases and a significant weight gain (median 7.75 kg, range 5 to 11). One patient developed a fatal intestinal T-cell lymphoma. CONCLUSIONS: In this case series, we have described several cases of CD in patients over the age of 60 with a varied spectrum of manifestations. We have also found a significant lag in diagnosis and treatment. We believe that it is important to promote the identification of CD as a possible culprit in varied clinical conditions in the elderly population.

Patients' views on optimal visit length in primary care.

Visit length (VL) has been decreasing over the last decades. Patients and physicians alike hold that this may have adverse affects on quality of care and patient-doctor rapport. Our aim was to study the optimal VL as viewed by the patients as well as possible related factors such as demographicparameters and patient satisfaction. By using surveys, we determined that patient satisfaction was highly correlated with VL and satisfaction from VL. Optimal VL as viewed by the patients was 15.4 minutes on average with considerable variation. Longer expected VL was associated with longer visits. Patient satisfaction from the visit was significantly decreased when expressed optimal VL exceeded the estimated duration of the visit. The surveys demonstrated that in young adults, satisfaction is highly correlated with VL. When asked, patients expect attainable VLs, which are on average only somewhat longer then the scheduled duration. We believe that patients should take part in decisions regarding visit scheduling. We suggest that individualization may contribute to the cost-effectiveness of physicians' time.

The pathophysiology of HCV induced B-cell clonal disorders.

Hepatitis C virus (HCV) has been shown in epidemiologic studies to be associated with immune system disorders. Primarily disorders that stem from B-cell regulatory control disturbance, such as mixed cryoglobulinemia (MC) and non-Hodgkin's lymphoma (NHL). The causative role of HCV in these disorders is supported by the response to anti-viral treatment. The understanding of the pathophysiological process leading from HCV infection to B-cell clonal expansion has improved significantly. Data supports an antigen-driven indirect stimulation of clonal expansion model, leading from oligoclonal to monoclonal expansion and in some instances to frank malignancy. HCV-E2 antigen has been suggested as a candidate antigen as well as NS3. Binding of the B-cell receptor by viral antigens coupled with direct binding of CD-81 by HCV-E2 has been shown to provide a strong proliferative signal. Additional regulatory elements are also affected in HCV-related B-cell clonal expansion, including the Fas and BLyS signaling mechanisms. Finally, genetic events such as bcl-2 rearrangement may also be involved in clonal expansion. In this review, evidence linking HCV with MC and NHL, as well as known events in the pathophysiological process are described.

Medex test, a novel modality for liver disease diagnosis: a pilot study.

BACKGROUND AND AIMS: Liver diseases are associated with significant morbidity and health- related expenditure. Although cost-effective treatments are available, the disease is often asymptomatic until late in its course. "Medex Test," is the noninvasive detection of liver abnormalities by the measurement of changes in electrical impedance of dermal zones. This method is based on neuroreflexology, a branch of complementary medicine. This study addressed 2 questions: can Medex Test detect liver disease, and can it measure the severity of a known liver disease. METHODS: This blinded case-control study included 2 parts. First, 113 patients with a known liver disease (hepatitis C, hepatitis B, and nonalcoholic fatty liver disease) and 85 controls with no known liver disease were evaluated by the Medex Test device. Second, necroinflammatory grading of biopsy results of 60 patients with chronic hepatitis C were compared with grade determined by Medex Test. RESULTS: Medex Test detected with high sensitivity (85%) and specificity (94.1%) the presence of liver disorders. The high rates were similar for the different disorders and were independent of age and sex. Additionally, Medex Test matched the biopsy pathologic grading of necroinflammation in 78% of patients. Positive predictive value was not affected by age and sex and was better for higher degree of necroinflammation. CONCLUSIONS: This pilot study demonstrated that Medex Test detects with high accuracy the presence of liver disorders and the necroinflammatory grade. This noninvasive, low cost test may in the future become an important tool in the diagnosis and management of liver disorders. We believe the further study of this novel method is warranted.