Postruminal choline supply during negative nutrient balance alters components of hepatic mTOR signaling and plasma amino acids in lactating Holstein cows.

Choline requirements for dairy cattle are unknown. However, enhanced postruminal supply of choline may increase flux through the methionine cycle to spare Met for other functions such as protein synthesis and phosphatidylcholine (PC) synthesis during periods of negative nutrient balance (NNB). The objective was to investigate the effects of postruminal choline supply during a feed restriction-induced NNB on hepatic abundance and phosphorylation of mTOR (mechanistic target of rapamycin)-related signaling proteins, hepatic lipidome and plasma AA. Ten primiparous rumen-cannulated Holstein cows (158 ± 24 DIM) were used in a replicated 5 × 5 Latin square design with 4 d of treatment and 10 d of recovery (14 d/period). Treatments were unrestricted intake with abomasal infusion of water, restricted intake (R; 60% of net energy for lactation requirements to induce NNB) with abomasal infusion of water (R0) or restriction plus abomasal infusion of 6.25, 12.5, or 25 g/d choline ion. Liver tissue was collected via biopsy on d 5 after infusions ended and used for Western blot analysis to measure proteins involved in mTOR signaling and untargeted lipidomics. Blood was collected on d 1 to 5 for plasma AA analysis. Statistical contrasts for protein and AA data were A0 versus R0 (CONT1), R0 versus the average of choline dose (CONT2) and tests of linear and quadratic effects of choline dose. Analysis of lipidomic data were performed with the web-based metabolomic processing tool MetaboAnalyst 5.0. Ratios of p-RPS6KB1:tRPS6KB1, p-EEF2:tEEF2, and p-EIF2:tEIF2 were greater with R (CONT1). Among those, supply of choline led to decreases in p-EEF2:tEEF2 (CONT2), p-EIF2:tEIF2 and tended to decrease p-EIF4BP1:tEIF4BP1. However, the effect was quadratic only for p-EEF2:tEEF2 and p-EIF2A:tEIF2A, reaching a nadir at 6.25 to 12.5 g/d choline ion. The ratio of p-RPS6KB1:tRPS6KB1 was not affected by supply of choline and was close to 2-fold greater at 25 g/d choline versus A0. Plasma Met concentration decreased with R (CONT1), but increased linearly with choline. Restriction also increased plasma 3-methyl-histidine (CONT1). The partial least squares discriminant analysis model of liver lipids distinguished treatments, with 13.4% of lipids being modified by treatment. One-way ANOVA identified 109 lipids with a false discovery rate ≤0.05. The largest group identified was PC species; all 35 detected decreased with R versus A0, but there were few differences among choline treatments. Overall, data suggested that dephosphorylation of EEF2 and EIF2A due to enhanced choline supply potentially helped maintain or increase protein synthesis during NNB. While activation of mTOR was not altered by choline, this idea of increased protein synthesis is partly supported by the increased circulating Met. However, enhanced postruminal choline had limited effects on the species of lipid produced during a period of NNB.

Identification of the First Gene Transfer Agent (GTA) Small Terminase in Rhodobacter capsulatus and Its Role in GTA Production and Packaging of DNA.

Genetic exchange mediated by viruses of bacteria (bacteriophages) is the primary driver of rapid bacterial evolution. The priority of viruses is usually to propagate themselves. Most bacteriophages use the small terminase protein to identify their own genome and direct its inclusion into phage capsids. Gene transfer agents (GTAs) are descended from bacteriophages, but they instead package fragments of the entire bacterial genome without preference for their own genes. GTAs do not selectively target specific DNA, and no GTA small terminases are known. Here, we identified the small terminase from the model Rhodobacter capsulatus GTA, which then allowed prediction of analogues in other species. We examined the role of the small terminase in GTA production and propose a structural basis for random DNA packaging.IMPORTANCE Random transfer of any and all genes between bacteria could be influential in the spread of virulence or antimicrobial resistance genes. Discovery of the true prevalence of GTAs in sequenced genomes is hampered by their apparent similarity to bacteriophages. Our data allowed the prediction of small terminases in diverse GTA producer species, and defining the characteristics of a "GTA-type" terminase could be an important step toward novel GTA identification. Importantly, the GTA small terminase shares many features with its phage counterpart. We propose that the GTA terminase complex could become a streamlined model system to answer fundamental questions about double-stranded DNA (dsDNA) packaging by viruses that have not been forthcoming to date.

The natural history of recovery of elbow flexion after obstetric brachial plexus injury managed without nerve repair.

In this study, we report the outcome for spontaneous recovery of elbow flexion in obstetric brachial plexus injury managed without nerve reconstruction. Excluding those with transient paralysis, our records revealed 152 children with obstetric brachial plexus injury born before our unit routinely offered brachial plexus reconstruction. Five had had nerve repairs. Of the remainder, only one patient had insufficient flexion to reach their mouth. Elbow flexion started to recover clinically at a mean age of 4 months for Narakas Group 1, 6 months for Group 2, 8 months for Group 3 and 12 months for Group 4. The mean active range of elbow flexion, in 44 cases, was 138°. The mean isometric elbow flexion strength, in 39 patients, was 63% (range 23%-100%) of the normal side. It appears to be rare for elbow flexion not to recover spontaneously, although recovery occurs later in more severe injuries. It is doubtful if nerve reconstruction can improve elbow flexion above the likely spontaneous recovery in babies with obstetric brachial plexus injuries. LEVEL OF EVIDENCE: II.

Digital histology quantification of intra-hepatic fat in patients undergoing liver resection.

BACKGROUND: High intra-hepatic fat (IHF) content is associated with insulin resistance, visceral adiposity, and increased morbidity and mortality following liver resection. However, in clinical practice, IHF is assessed indirectly by pre-operative imaging [for example, chemical-shift magnetic resonance (CS-MR)]. We used the opportunity in patients undergoing liver resection to quantify IHF by digital histology (D-IHF) and relate this to CT-derived anthropometrics, insulin-related serum biomarkers, and IHF estimated by CS-MR. METHODS: A reproducible method for quantification of D-IHF using 7 histology slides (inter- and intra-rater concordance: 0.97 and 0.98) was developed. In 35 patients undergoing resection for colorectal cancer metastases, we measured: CT-derived subcutaneous and visceral adipose tissue volumes, Homeostasis Model Assessment of Insulin Resistance (HOMA-IR), fasting serum adiponectin, leptin and fetuin-A. We estimated relative IHF using CS-MR and developed prediction models for IHF using a factor-clustered approach. RESULTS: The multivariate linear regression models showed that D-IHF was best predicted by HOMA-IR (Beta coefficient(per doubling): 2.410, 95% CI: 1.093, 5.313) and adiponectin (ß(per doubling): 0.197, 95% CI: 0.058, 0.667), but not by anthropometrics. MR-derived IHF correlated with D-IHF (rho: 0.626; p = 0.0001), but levels of agreement deviated in upper range values (CS-MR over-estimated IHF: regression versus zero, p = 0.009); this could be adjusted for by a correction factor (CF: 0.7816). CONCLUSIONS: Our findings show IHF is associated with measures of insulin resistance, but not measures of visceral adiposity. CS-MR over-estimated IHF in the upper range. Larger studies are indicated to test whether a correction of imaging-derived IHF estimates is valid.

Liver resection rate following downsizing chemotherapy with cetuximab in metastatic colorectal cancer: UK retrospective observational study.

AIMS: The high objective response rate to cetuximab along with chemotherapy in patients with colorectal liver metastases makes it an effective downsizing protocol to facilitate surgery in those with initially unresectable disease. Adoption of this strategy has been variable in the UK. A retrospective observational study was conducted in 7 UK specialist liver surgical centres to describe the liver resection rate following a downsizing protocol of cetuximab and chemotherapy and to evaluate the quality and efficiency of processes by which the treatment was provided. METHODS: Data were collected in 2012 by reviewing medical records of patients with colorectal metastases confined to the liver, defined as unresectable without downsizing therapy at first review by a specialist Multi Disciplinary Team (MDT). RESULTS: Sixty patients were included; 29 (48%) underwent liver resection following cetuximab and chemotherapy. Of the 29, 17 (59% or 28% of all patients) achieved R0 resection and 7 (24% or 12% of all patients) R1 resection. All treated patients were KRAS wild-type. CONCLUSION: In specialist liver surgical centres, where patients are evaluated for liver resection, optimal management by MDT using KRAS testing, cetuximab and chemotherapy results in a 28% R0 resection rate in patients with initially unresectable colorectal cancer liver metastases.

A comparison of diagnostic imaging modalities for colorectal liver metastases.

AIMS: The aims of this study were to compare the diagnostic performance of CT scan, MR liver, PET-CT and intra-operative ultrasound (IOUS) for the detection of liver metastases against the histopathological findings, and to compare PET-CT with CT for the detection of distant disease in metastatic colorectal cancer patients eligible for surgical treatment. METHODS: A prospective study was performed that measured concordance between the number and stage of metastatic lesions identified with various preoperative imaging modalities and histology of patients undergoing surgical treatment for CRLM. RESULTS: Compared with histopathology, concordance for the number of metastatic liver lesions was moderate for CT scan (K = 0.477, 95% CI: 0.28-0.66), moderate for MR scan (K = 0.574, 95% CI: 0.39-0.75), good for FDG PET-CT (K = 0.703, 95% CI: 0.52-0.87) and very good for IOUS (K = 0.904, 95% CI: 0.81-0.99). Additional CRLM were identified intraoperatively in six patients (9.1%) with IOUS and in 7.5% of the cases surgical strategy was changed according to the new intraoperative findings. The diagnosis of intra abdominal lymph node metastatic disease was made with PET-CT only in nine patients (13.6%) DISCUSSION: Our study supports the recent recommendations of the Oncosurg Multidisciplinary International Consensus regarding the importance of high quality CT and MR in the staging of CRLM but provides further evidence for the added value of PET-CT, especially in detecting extrahepatic intra-abdominal metastatic disease that may be amenable to potentially curative resection. Despite these advances in preoperative staging, there still remains a role for IOUS in detecting additional metastases at the time of surgery.

Management strategies for shoulder reconstruction in obstetric brachial plexus injury with special reference to loss of internal rotation after surgery.

We assessed the outcomes in 24 of 25 children (mean age 3 years) who had soft-tissue shoulder reconstruction procedures for obstetric brachial plexus injuries. All had latissimus dorsi and teres major transfers to strengthen external rotation combined with subscapularis lengthening, reduction of posterior dislocation or subluxation, and humeral osteotomy, if necessary. At a mean follow-up of 3.8 years, the mean Mallet score improved from 12.3 preoperatively to 17.4 postoperatively, active external rotation improved from 21° to 32°, and passive external rotation improved from 26° to 67°. The shoulder remained in joint in all patients. Active internal rotation deteriorated in nine shoulders, two requiring secondary internal rotation osteotomy. Combined soft tissue rebalancing and bony procedures gives reliable shoulder function improvement. Care is required regarding the effect on internal rotation.

Intestinal ciprofloxacin efflux: the role of breast cancer resistance protein (ABCG2).

Intestinal secretory movement of the fluoroquinolone antibiotic, ciprofloxacin, may limit its oral bioavailability. Active ATP-binding cassette (ABC) transporters such as breast cancer resistance protein (BCRP) have been implicated in ciprofloxacin transport. The aim of this study was to test the hypothesis that BCRP alone mediates intestinal ciprofloxacin secretion. The involvement of ABC transport proteins in ciprofloxacin secretory flux was investigated with the combined use of transfected cell lines [bcrp1/BCRP-Madin-Darby canine kidney II (MDCKII) and multidrug resistance-related protein 4 (MRP4)-human embryonic kidney (HEK) 293] and human intestinal Caco-2 cells, combined with pharmacological inhibition using 3-(6-isobutyl-9-methoxy-1,4-dioxo-1,2,3,4,6, 7,12,12a-octahydropyrazino[1',2':1,6]pyrido[3,4-b]indol-3-yl)-propionic acid tert-butyl ester (Ko143), cyclosporine, 3-[[3-[2-(7-chloroquinolin-2-yl)vinyl]phenyl]-(2-dimethylcarbamoylethylsulfanyl)methylsulfanyl] propionic acid (MK571), and verapamil as ABC-selective inhibitors. In addition, the regional variation in secretory capacity was investigated using male Han Wistar rat intestine mounted in Ussing chambers, and the first indicative measurements of ciprofloxacin transport by ex vivo human jejunum were made. Active, Ko143-sensitive ciprofloxacin secretion was observed in bcrp1-MDCKII cell layers, but in low-passage (BCRP-expressing) Caco-2 cell layers only a 54% fraction was Ko143-sensitive. Ciprofloxacin accumulation was lower in MRP4-HEK293 cells than in the parent line, indicating that ciprofloxacin is also a substrate for this transporter. Ciprofloxacin secretion by Caco-2 cell layers was not inhibited by MK571. Secretory flux showed marked regional variability in the rat intestine, increasing from the duodenum to peak in the ileum. Ciprofloxacin secretion was present in human jejunum and was reduced by Ko143 but showed marked interindividual variability. Ciprofloxacin is a substrate for human and rodent BCRP. An additional pathway for ciprofloxacin secretion exists in Caco-2 cells, which is unlikely to be MRP(4)-mediated. BCRP is likely to be the dominant transport mechanism for ciprofloxacin efflux in both rat and human jejunum.

Evaluation of surgical outcome scores according to ISGPS definitions in patients undergoing pancreatic resection.

AIMS: To evaluate the role of the Physiological and Operative Severity Score for the enUmeration of Mortality and Morbidity (POSSUM), the Portsmouth variant (p-POSSUM) and the Glasgow Prognostic Score (GPS) in predicting outcome after pancreatic surgery with reference to the International Study Group of Pancreatic Surgery (ISGPS) definitions of post-pancreatectomy complications. METHODS: All consecutive patients undergoing major pancreatic resection over a 32- month period were included. POSSUM, p-POSSUM and GPS score were calculated for each patient and correlated against the observed mortality and morbidity using the ISGPS definitions. RESULTS: The observed:expected ratios for POSSUM mortality, POSSUM morbidity and p-POSSUM mortality were 0.24 (p < 0.0001), 0.86 (p < 0.0001) and 0.79 (p = 0.09), respectively. POSSUM had a 'poor fit' with respect to predicting morbidity (χ(2) = 16.4, 8 d.f., p = 0.04). Multifactorial regression analysis revealed the GPS as an independent predictor of post-operative outcome (GPS 1, p = 0.03, OR 2.99, 95% CI 1.4-7.9, and GPS 2, p = 0.02, OR 4.3, 95% CI 1.8-15.5). CONCLUSION: POSSUM has a limited role as an outcome score in pancreatic resection. The GPS may be a novel alternative to POSSUM as a pre-operative predictor of outcome.

Outcome of late-onset Perthes' disease using four different treatment modalities.

PURPOSE: The outcome of Perthes' disease in children over 8 years tends to be poor. It is unclear whether any treatment modality alters the natural history. This study compares the results of four treatment modalities for this group of patients. METHODS: A retrospective review was performed of prospectively collected data for 44 children (48 hips) with Catterall grade 2, 3 or 4 Perthes' disease with onset age 8 years or older followed to maturity. Patients were divided into four groups (a no-treatment group and three interventional groups). The interventional groups were demographically similar but the untreated patients were older and had poorer indices for most modalities. RESULTS: Overall for all treatment modalities only 19% had a satisfactory Stulberg grade II outcome. Poorer outcomes (as assessed by center-edge angle and percentage femoral head coverage) were associated with increasing age, greater initial head deformity, and more head involvement. Initial head deformity did not remodel for any group and progressed despite plaster treatment or varus osteotomy but not after acetabular augmentation. Acetabular augmentation gave better outcomes for C/B ratio and center-edge angle. CONCLUSIONS: Whatever the treatment, the outcome is poorer with increasing age. No treatment guarantees a good result for significant head involvement or initial deformity but acetabular augmentation improves C/B ratio and center-edge angle and prevents progressive femoral head deformity compared with no treatment, varus osteotomy, and plaster treatment.

Legg-Calvé-Perthes disease affecting children less than eight years of age: a paired outcome study.

There is controversy as to whether operative treatment in Legg-Calvé-Perthes (LCP) disease improves the outcome obtained by conservative treatment. This article describes a paired study of mature patients, diagnosed with LCP disease before the age of 8 years, to compare the radiological results after conservative or surgical management. Patients were paired with strict matching of gender, body mass index, age at onset, disease stage at the first visit, necrotic area, and radiological at-risk signs. Each pair was assessed by comparing the values of five radiological measurements. Fourteen pairs (28 hips) fitted the criteria. The only radiological measure which showed a statistically better result in the surgical group was femoral head sphericity as measured by Mose's method. However, there was no difference between the two groups for Stulberg class. Our study supports the hypothesis that for hips affected with LCP disease under the age of 8 years, surgery does not improve the outcome.

Outcome of lateral humeral condylar mass fractures in children associated with elbow dislocation or olecranon fracture.

Of 2,502 elbow/humeral injuries treated at our department between 1990 and 2005, we identified a cohort of 20 lateral condylar mass (LCM) fractures of the humerus in children associated with elbow dislocation (n = 12; mean age 8.2 years) or with olecranon fracture (n = 8; mean age 4.1 years). Eight patients with undisplaced fracture pattern were treated conservatively yielding a satisfactory outcome. Good to excellent results were obtained in the majority (85%). Overall, the result was poor in three patients (15%; 25% of the operated cohort) due to terminal 20-30 degrees loss of extension. There was no obvious difference in the outcome between the isolated displaced LCM fractures described in the literature and this cohort. Testing of elbow stability by examination under anaesthesia is stressed. Undisplaced fracture patterns need to be closely observed. Parents should be warned about the likelihood of some degree of unfavourable outcome in the displaced LCM fractures with associated elbow injuries.

Chemoradiotherapy for locally advanced pancreatic cancer: a radiotherapy dose escalation and organ motion study.

AIMS: To determine the efficacy of radiation dose escalation and to examine organ motion during conformal radiotherapy for locally advanced pancreatic cancer. MATERIALS AND METHODS: Thirty-nine patients who were consecutively treated with chemoradiotherapy were studied. Fifteen patients, treated from 1993 to 1997, received 50 Gy in 20 fractions (group I). Twenty-four patients, treated from 1997 to 2003, received an escalated dose of 55 Gy in 25 fractions (group II). Intra-fraction pancreatic tumour motion was assessed in three patients using megavoltage movies during radiation delivery to track implanted radio-opaque markers. RESULTS: Improved survival rates were seen in latterly treated group II patients (P=0.083), who received escalated radiotherapy to smaller treatment volumes due to advances in verification. Worse toxicity effects (World Health Organization grade 3-4) were reported by some patients (<10%), but treatment compliance was similar in both groups, indicating equivalent tolerance. Substantial intra-fraction tumour displacement due to respiratory motion was observed: this was greatest in the superior/inferior (mean=6.6 mm) and anterior/posterior (mean=4.75 mm) directions. Lateral displacements were small (<2 mm). CONCLUSIONS: Dose escalation is feasible in pancreatic cancer, particularly when combined with a reduction in irradiated volume, and enhanced efficacy is indicated. Large, globally applied margins to compensate for pancreatic tumour motion during radiotherapy may be inappropriate. Strategies to reduce respiratory motion, and/or the application of image-guided techniques that incorporate individual patients' respiratory motion into radiotherapy planning and delivery, will probably improve pancreatic radiotherapy.

Lymphatic vessel density, microvessel density and lymphangiogenic growth factor expression in colorectal cancer.

OBJECTIVE: Microvessel density (MVD) has been studied as a prognostic marker in human cancers. Quantification of lymphatic vessel density (LVD) is now possible by using new antibodies. Expression of the lymphangiogenic growth factors, VEGF-C and VEGF-D, is associated with poorer clinicopathological outcomes in various tumours. The aim of this study was to quantify LVD and MVD in colorectal cancer, determine the relationship between LVD, MVD and clinicopathological variables and examine the relationship between LVD and tumour expression of VEGF-C and VEGF-D. METHOD: Thirty primary colorectal cancers were immunostained for CD34, lymph vessel endothelial hyaluronan receptor-1 (LYVE-1), VEGF-A and VEGF-D using standard techniques. LVD and MVD were determined by Chalkley grid counting. Tumours were assessed for the presence or absence of LYVE-1 positive lymphatics at different areas within the tumour and the tumour was scored for VEGF-C and VEGF-D immunostaining intensity at the invading tumour edge. Non-parametric tests were used for statistical analysis and a P-value of <0.05 was taken as significant. RESULTS: Lymph vessel endothelial hyaluronan receptor-1 was an excellent lymphatic vessel marker. Within normal bowel wall, lymphatic vessels were found rarely in the superficial colonic mucosa, but were numerous in the submucosa and muscularis propria. In the majority of tumours, lymphatic vessels were located in the peri-tumoural area, intra-tumoural vessels were sparse and tended to be narrow with closed lumina. At the invading tumour edge, VEGF-C expression was higher (P = 0.028) and VEGF-D expression lower (P = 0.011), in tumours in which lymphatic vessels were present. No significant differences between LVD and any clinicopathological variable or route of metastasis were identified. CONCLUSION: Lymphatic vessel density and MVD can be quantified in colorectal carcinoma using immunohistochemical techniques. The balance between expression of VEGF-C and VEGF-D at the invading tumour edge may enhance lymphatic metastasis, by promoting tumour lymphangiogenesis or by activation of pre-existing lymphatic vessels. No relationship was identified between LVD and clinicopathological variables.

Impact of laparotomy and liver resection on the peritoneal concentrations of fibroblast growth factor 2, vascular endothelial growth factor and hepatocyte growth factor.

PURPOSE: Some data have suggested that major surgery is associated with the post-operative growth of residual tumour masses but the mechanism of this is unknown. This study was designed to determine the relationship between intraperitoneal (IP) cytokine levels, and laparotomy in benign and malignant settings. METHODS: Intraperitoneal fluid specimens were obtained at the start and at the end of laparotomy in patients with benign conditions (n=10) and in others undergoing resection of hepatic metastases from colorectal cancer (n=10). Using ELISA the concentration of the angiogenic cytokines, HGF, VEGF-A, VEGF-C, VEGF-D and FGF-2 was determined. RESULTS: The data show that in 16 of 20 patients there was a significant increase (P=0.006) in the IP concentration of hepatocyte growth factor (HGF) but not in the other growth factors by the end of the operation. The mean increase in HGF concentration was 821.5 pg/ml (95% CI: 11.0-6,426.0). Neither the groups (malignant and non-malignant) nor the length of operation correlated with greater or lesser increases in HGF. CONCLUSION: The observation that the increase in HGF occurred in both the cancer and non-cancer groups suggests that it is the surgery rather than the disease that is associated with the increased cytokine concentration. As HGF is a potent endothelial, epithelial and mesenchymal mitogen the data highlight HGF as a potential target for anti-cancer treatments in the peri-operative period. However, investigators should closely monitor wound healing as this may be compromised by this new class of drugs.

A profile of Perthes' disease in Greater Glasgow: is there an association with deprivation?

It has been reported that there is an association between Perthes' disease and poverty. We examined the demographic data of a group of 240 children (263 hips) who presented with Perthes' disease in Greater Glasgow, where the mean deprivation scores are substantially greater than in the rest of Scotland, to see if this association applied and whether other clues to the aetiology of Perthes' disease could be found. There were 197 boys and 43 girls; 39 (16.25%) had a family history of Perthes' disease. Bone age in this series was heavily skewed towards the lower percentiles. The mean number of siblings was 1.9, with 31 (12.9%) being an only child. Maternal age at the birth of the first child showed no preponderance of older mothers. Maternal smoking during and after pregnancy was noted in 132 (55%), which compared with the 52% reported in the population of Greater Glasgow in general. Of the children in our series, 60 (25%) were in social class IV and V. However, this applies to more than half of the population of Greater Glasgow. There was no significant evidence of a preponderance of Perthes' disease in the most deprived groups. The aetiology of Perthes' disease is likely to be multifactorial and may include a genetic or deprivation influence resulting in delayed bone age.

Treatment of inoperable hepatocellular carcinoma with pegylated liposomal doxorubicin (PLD): results of a phase II study.

Monthly intravenous pegylated liposomal doxorubicin (PLD) 50 mg m(-2), although well tolerated, showed almost no activity in this phase II study of 16 patients with advanced hepatocellular carcinoma with a response rate of 0%, stable disease 19%, median time to progression of 2.4 months, 1-year survival of 25% and median survival of 6.5 months.

Effect of patient, operative and isolation factors on subsequent yield and viability of human hepatocytes for research use.

It is widely accepted that the model of choice for pharmacotoxicological studies are human hepatocytes. There is therefore a demand for these cells, but quality must be maintained for their widespread use. We present a retrospective review of the isolation of hepatocytes from both surgically resected tissue and livers rejected for transplantation, and evaluated patient, operative and isolation variables to ascertain which may affect the viability and yield of cells. Seven clinically rejected whole livers and 60 surgically resected specimens (from two distinct operating centres) were isolated. For surgically resected tissue we found that decreasing age, securing the perfusing cannulae with suture rather than reforming Glissons capsule with glue and steatotic livers improved viability. No significant correlation could be found with pre-operative blood results, disease, type of operation, presence or absence of Pringle manoeuvre, weight of tissue isolated, time of digestion with collagenase and cold ischaemic time. There was a reduction in mean yield and viability when hepatocyte isolations were performed in livers rejected for transplant, compared to surgically resected tissue although this did not reach significance. Human hepatocytes can be successfully and consistently isolated from surgically resected tissue and appear to be superior to those isolated from rejected for transplant livers. From our study, there are few parameters that significantly affect the quality of isolated hepatocytes, which increases the possible pool of tissue that hepatocytes can be isolated from.