Plaque burden estimated from optical coherence tomography with deep learning: In vivo validation using co-registered intravascular ultrasound.

OBJECTIVES: The objective of the present study was to compare plaque burden (PB) calculated from optical coherence tomography (OCT) using deep learning (DL) with PB derived from co-registered intravascular ultrasound (IVUS). BACKGROUND: A DL algorithm was developed for automated plaque characterization and PB quantification from OCT images. However, the performance of this algorithm for PB quantification has not been validated. METHODS: Five-year follow-up OCT and IVUS images from 15 patients implanted with bioresorbable vascular scaffold (BVS) at baseline were analyzed. Precise co-registration for 72 anatomical slices was achieved utilizing unique BVS radiopaque markers. PB derived from OCT DL and IVUS were compared. OCT cross-sections were divided into four subgroups with different media visibility level. The impact of media visibility on the numerical difference between OCT-derived and IVUS-derived PB was investigated. The stent sizes selected by OCT DL and IVUS were compared. RESULTS: Sixty-four paired OCT and IVUS cross-sections were compared. OCT DL showed good concordance with IVUS for PB assessment (ICC = 0.81, difference = -3.53 ± 6.17%, p < 0.001). The numerical difference between OCT DL-derived PB and IVUS-derived PB was not substantially impacted by missing segments of media visualization (p = 0.21). OCT DL showed a diagnostic accuracy of 92% in identifying PB > 65%. The stent sizes selected by OCT DL were smaller compared to the ones selected by IVUS (difference = 0.30 ± 0.34 mm, p < 0.001). CONCLUSIONS: The DL algorithm provides a feasible and reliable method for automated PB estimation from OCT, irrespective of media visibility. OCT DL showed good diagnostic accuracy in identifying PB > 65%, revealing its potential to complement conventional OCT imaging.

Tissue-based next generation sequencing: application in a universal healthcare system.

In the context of solid tumours, the evolution of cancer therapies to more targeted and nuanced approaches has led to the impetus for personalised medicine. The targets for these therapies are largely based on the driving genetic mutations of the tumours. To track these multiple driving mutations the use of next generation sequencing (NGS) coupled with a morphomolecular approach to tumours, has the potential to deliver on the promises of personalised medicine. A review of NGS and its application in a universal healthcare (UHC) setting is undertaken as the technology has a wide appeal and utility in diagnostic, clinical trial and research paradigms. Furthermore, we suggest that these can be accommodated with a unified integromic approach. Challenges remain in bringing NGS to routine clinical use and these include validation, handling of the large amounts of information flow and production of a clinically useful report. These challenges are particularly acute in the setting of UHC where tests are not reimbursed and there are finite resources available. It is our opinion that the challenges faced in applying NGS in a UHC setting are surmountable and we outline our approach for its routine application in diagnostic, clinical trial and research paradigms.

Back to the future: routine morphological assessment of the tumour microenvironment is prognostic in stage II/III colon cancer in a large population-based study.

AIMS: Both morphological and molecular approaches have highlighted the biological and prognostic importance of the tumour microenvironment in colorectal cancer (CRC). Despite this, microscopic assessment of the tumour microenvironment has not been adopted into routine practice. The study aim was to identify those tumour microenvironmental features that are most likely to provide prognostic information and be feasible to use in routine pathology reporting practice. METHODS AND RESULTS: On the basis of existing evidence, we selected specific morphological features relating to peritumoral inflammatory and stromal responses, agreed criteria for scoring, and assessed these in representative haematoxylin and eosin (H&E)-stained whole tumour sections from a population-based cohort of 445 stage II/III colon cancer cases. Moderate/severe peritumoral diffuse lymphoid inflammation and Crohn's disease-like reaction were associated with significantly reduced risks of CRC-specific death [adjusted hazard ratio (HR) 0.48, 95% confidence interval (CI) 0.31-0.76, and HR 0.60, 95% CI 0.42-0.84, respectively]. The presence of >50% tumour stromal percentage, as assessed by global evaluation of tumour area, was associated with a significantly increased risk of CRC-specific death (HR 1.60 95% CI 1.06-2.41). A composite 'fibroinflammatory score' (0-3), combining dichotomized scores of these three features, showed a highly significant association with survival outcomes. Those with a score of ≥2 had an almost 2.5-fold increased risk of CRC-specific death (HR 2.44, 95% CI 1.56-3.81) as compared with those scoring zero. These associations were stronger in microsatellite instability (MSI)-high tumours, potentially identifying a subset of MSI-high colon cancers that lack characteristic morphological features and have an associated worse prognosis. CONCLUSIONS: In summary, reporting on H&E staining of selected microscopic features of the tumour microenvironment, independently or in combination, offers valuable prognostic information in stage II/III colon cancer, and may allow morphological correlation with developing molecular classifications of prognostic and predictive relevance.

Human umbilical cord-derived mesenchymal stromal cells improve myocardial fibrosis and restore miRNA-133a expression in diabetic cardiomyopathy.

BACKGROUND: Diabetic cardiomyopathy (DCM) is a serious health-threatening complication of diabetes mellitus characterized by myocardial fibrosis and abnormal cardiac function. Human umbilical cord mesenchymal stromal cells (hUC-MSCs) are a potential therapeutic tool for DCM and myocardial fibrosis via mechanisms such as the regulation of microRNA (miRNA) expression and inflammation. It remains unclear, however, whether hUC-MSC therapy has beneficial effects on cardiac function following different durations of diabetes and which mechanistic aspects of DCM are modulated by hUC-MSC administration at different stages of its development. This study aimed to investigate the therapeutic effects of intravenous administration of hUC-MSCs on DCM following different durations of hyperglycemia in an experimental male model of diabetes and to determine the effects on expression of candidate miRNAs, target mRNA and inflammatory mediators. METHODS: A male mouse model of diabetes was induced by multiple low-dose streptozotocin injections. The effects on severity of DCM of intravenous injections of hUC-MSCs and saline two weeks previously were compared at 10 and 18 weeks after diabetes induction. At both time-points, biochemical assays, echocardiography, histopathology, polymerase chain reaction (PCR), immunohistochemistry and enzyme-linked immunosorbent assays (ELISA) were used to analyze blood glucose, body weight, cardiac structure and function, degree of myocardial fibrosis and expression of fibrosis-related mRNA, miRNA and inflammatory mediators. RESULTS: Saline-treated diabetic male mice had impaired cardiac function and increased cardiac fibrosis after 10 and 18 weeks of diabetes. At both time-points, cardiac dysfunction and fibrosis were improved in hUC-MSC-treated mice. Pro-fibrotic indicators (α-SMA, collagen I, collagen III, Smad3, Smad4) were reduced and anti-fibrotic mediators (FGF-1, miRNA-133a) were increased in hearts of diabetic animals receiving hUC-MSCs compared to saline. Increased blood levels of pro-inflammatory cytokines (IL-6, TNF, IL-1ß) and increased cardiac expression of IL-6 were also observed in saline-treated mice and were reduced by hUC-MSCs at both time-points, but to a lesser degree at 18 weeks. CONCLUSION: Intravenous injection of hUC-MSCs ameliorated key functional and structural features of DCM in male mice with diabetes of shorter and longer duration. Mechanistically, these effects were associated with restoration of intra-myocardial expression of miRNA-133a and its target mRNA COL1AI as well as suppression of systemic and localized inflammatory mediators.

Amelioration of diabetic nephropathy in mice by a single intravenous injection of human mesenchymal stromal cells at early and later disease stages is associated with restoration of autophagy.

BACKGROUND AND AIMS: Mesenchymal stromal cells (MSCs) a potentially effective disease-modulating therapy for diabetic nephropathy (DN) but their clinical translation has been hampered by incomplete understanding of the optimal timing of administration and in vivo mechanisms of action. This study aimed to elucidate the reno-protective potency and associated mechanisms of single intravenous injections of human umbilical cord-derived MSCs (hUC-MSCs) following shorter and longer durations of diabetes. METHODS: A streptozotocin (STZ)-induced model of diabetes and DN was established in C57BL/6 mice. In groups of diabetic animals, human (h)UC-MSCs or vehicle were injected intravenously at 8 or 16 weeks after STZ along with vehicle-injected non-diabetic animals. Diabetes-related kidney abnormalities was analyzed 2 weeks later by urine and serum biochemical assays, histology, transmission electron microscopy and immunohistochemistry. Serum concentrations of pro-inflammatory and pro-fibrotic cytokines were quantified by ELISA. The expression of autophagy-related proteins within the renal cortices was investigated by immunoblotting. Bio-distribution of hUC-MSCs in kidney and other organs was evaluated in diabetic mice by injection of fluorescent-labelled cells. RESULTS: Compared to non-diabetic controls, diabetic mice had increases in urine albumin creatinine ratio (uACR), mesangial matrix deposition, podocyte foot process effacement, glomerular basement membrane thickening and interstitial fibrosis as well as reduced podocyte numbers at both 10 and 18 weeks after STZ. Early (8 weeks) hUC-MSC injection was associated with reduced uACR and improvements in multiple glomerular and renal interstitial abnormalities as well as reduced serum IL-6, TNF-α, and TGF-ß1 compared to vehicle-injected animals. Later (16 weeks) hUC-MSC injection also resulted in reduction of diabetes-associated renal abnormalities and serum TGF-ß1 but not of serum IL-6 and TNF-α. At both time-points, the kidneys of vehicle-injected diabetic mice had higher ratio of p-mTOR to mTOR, increased abundance of p62, lower abundance of ULK1 and Atg12, and reduced ratio of LC3B to LC3A compared to non-diabetic animals, consistent with diabetes-associated suppression of autophagy. These changes were largely reversed in the kidneys of hUC-MSC-injected mice. In contrast, neither early nor later hUC-MSC injection had effects on blood glucose and body weight of diabetic animals. Small numbers of CM-Dil-labeled hUC-MSCs remained detectable in kidneys, lungs and liver of diabetic mice at 14 days after intravenous injection. CONCLUSIONS: Single intravenous injections of hUC-MSCs ameliorated glomerular abnormalities and interstitial fibrosis in a mouse model of STZ-induced diabetes without affecting hyperglycemia, whether administered at relatively short or longer duration of diabetes. At both time-points, the reno-protective effects of hUC-MSCs were associated with reduced circulating TGF-ß1 and restoration of intra-renal autophagy.

The strategic use of biopsy in the diagnosis of coeliac disease in adults.

BACKGROUND: The paediatric guidelines support the use of the 'No Biopsy Approach' in the diagnosis of coeliac disease (CD). We aimed to determine the correlation between anti tissue transglutaminase (anti-TTG serology) ≥10 times the upper limit of normal (ULN), using the Celikey ® ELiA assay and histological findings. Our secondary aim was to determine the safety of this approach in our centre. METHODS: A retrospective analysis of adult patients referred to a tertiary referral centre with raised anti-TTG titres and/or histological changes of coeliac on D2 biopsies between 2014 - 2019. Excluded patients were those who did not have a biopsy performed, or whose biopsy was unavailable for review, selective IgA deficiency, and gluten elimination prior to biopsy. Biopsies were classified according to Marsh, by two independent pathologists, blinded to the anti-TTG titre. RESULTS: 164 patients had positive anti-TTG serology and duodenal biopsy in our centre prior to starting a gluten free diet (GFD) in the period 2014 - 2019. Of these 164 patients (median age 40yrs, 62% female), 68 (33%) had an anti-TTG titre ≥10 x ULN, 99% of which had a Marsh grading ≥ 3 and 1% had a Marsh of 2 on biopsy. 91% had either a normal index gastroscopy or findings of mild gastritis/oesophagitis. CONCLUSIONS: We found a 98.5% positive predictive value (PPV) of determining CD (i.e., Marsh ≥ 3) in those with an anti-TTG ≥10 x ULN. In those with moderate to high-risk clinical suspicion of CD we propose that duodenal biopsy is unnecessary for diagnosis.

Targeting stromal cell sialylation reverses T cell-mediated immunosuppression in the tumor microenvironment.

Immunosuppressive tumor microenvironments (TMEs) reduce the effectiveness of immune responses in cancer. Mesenchymal stromal cells (MSCs), precursors to cancer-associated fibroblasts (CAFs), promote tumor progression by enhancing immune cell suppression in colorectal cancer (CRC). Hyper-sialylation of glycans promotes immune evasion in cancer through binding of sialic acids to their receptors, Siglecs, expressed on immune cells, which results in inhibition of effector functions. The role of sialylation in shaping MSC/CAF immunosuppression in the TME is not well characterized. In this study, we show that tumor-conditioned stromal cells have increased sialyltransferase expression, α2,3/6-linked sialic acid, and Siglec ligands. Tumor-conditioned stromal cells and CAFs induce exhausted immunomodulatory CD8+ PD1+ and CD8+ Siglec-7+/Siglec-9+ T cell phenotypes. In vivo, targeting stromal cell sialylation reverses stromal cell-mediated immunosuppression, as shown by infiltration of CD25 and granzyme B-expressing CD8+ T cells in the tumor and draining lymph node. Targeting stromal cell sialylation may overcome immunosuppression in the CRC TME.

A unique cause of a rare disorder, unilateral macromastia due to lymphangiomatosis of the breast: a case report.

Macromastia and in particular unilateral macromastia is a rare clinical entity. It relates to massive enlargement of the breast in non-obese women. This case report describes an initially mild unilateral asymmetry occurring nine months postpartum in a 33-year-old female. However, following her second pregnancy within 12 months, her left breast became severely enlarged. This did not improve on delivery. No discrete lesion was seen on imaging and no significant abnormalities were seen in her blood chemistry. Surgical treatment was a mammoplasty and 580 g of mammary tissue was removed. Grossly, there was spongiform subcutaneous tissue with diffuse extension. On histology, this consisted of a highly complex and diffuse pattern of infiltration and of cavernous empty channels lined by a delicate attenuated endothelium which was CD34 and D2-40 positive. The appearances were consistent with lymphangiomatosis, more commonly encountered in the limbs, heretofore. Lymphangiomatosis has not been previously described in breast tissue and only a single case report exists for such a lesion in axillary tissue. Treatment of such lesions in the periphery by surgical excision is very difficult and excision without being radical can be impossible. Follow up of our patient, shows no evidence of recurrence in this patient. Our case report describes the clinicopathological features, differential diagnosis to be considered and treatment, in addition to reviewing the relevant literature.

Case report of the duplex vermiform appendix: A rare presentation in an elderly patient.

INTRODUCTION AND IMPORTANCE: Duplex appendix is a rare anatomical entity with incidence rate of 0.004 and 0.009%. Diagnosis is often missed despite growth in radiological investigations. Missed appendiceal anomalies can lead to undesirable medicolegal implications. CASE PRESENTATION: Here we discuss a case of a 76-year-old-male who initially presented to his primary care physician with right-sided abdominal pain for several weeks. A colonoscopy was performed and demonstrated a lesion arising from the appendicular orifice. The patient underwent staging imaging including Computerised Tomography of the abdomen and pelvis which demonstrated a dilated appendix. The patient underwent a laparoscopic right hemicolectomy. He made an uneventful recovery post-operatively and at his follow-up review at 4 weeks and 2 months. DISCUSSION: While duplex appendix has been reported in the literature, to our knowledge this is the first case report to describe duplicated appendix presenting as a colonic mass in an elderly patient. CONCLUSION: Intra-operative examination of the cecum is paramount to rule out appendiceal anomalies and prevent medicolegal complications.

Inflammation-specific targeted carriers for local drug delivery to inflammatory bowel disease.

Delivering drugs directly to the inflamed intestinal sites to treat inflammatory bowel disease (IBD), particularly Crohn's and ulcerative colitis, is highly challenging. Recent advances in colitis therapy medications are expanding opportunities for improving local on-site drug availability by minimising the associated systemic side-effects. Drug delivery with targeted carrier systems has shown the potential to increase site-specificity, stability, and therapeutic efficacy. Herein, we report the development of a strong anionic charged inflammation targeted nanocarriers (IT-NCs) loaded with an immunosuppressant model drug. This system showed preferential adhesion on a charge-modified surface in vitro, and in both dextran sulfate sodium (DSS) and TNBS colitis mice in vivo models. IT-NCs showed improved colitis phenotype therapeutic efficacy in both animal models compared to free drug. Furthermore, ex vivo study of colon tissue biopsies from patients with colitis revealed that IT-NCs adhered preferentially to inflamed biopsies compared to normal. Together, our results suggest that IT-NCs have promising therapeutic potential as delivery carriers' in colitis management.

Comparison of Single and Repeated Dosing of Anti-Inflammatory Human Umbilical Cord Mesenchymal Stromal Cells in a Mouse Model of Polymicrobial Sepsis.

Mesenchymal stromal cells (MSCs) ameliorate pre-clinical sepsis and sepsis-associated acute kidney injury (SA-AKI) but clinical trials of single-dose MSCs have not indicated robust efficacy. This study investigated immunomodulatory effects of a novel MSC product (CD362-selected human umbilical cord-derived MSCs [hUC-MSCs]) in mouse endotoxemia and polymicrobial sepsis models. Initially, mice received intra-peritoneal (i.p.) lipopolysaccharide (LPS) followed by single i.p. doses of hUC-MSCs or vehicle. Next, mice underwent cecal ligation and puncture (CLP) followed by intravenous (i.v.) doses of hUC-MSCs at 4 h or 4 and 28 h. Analyses included serum/plasma assays of biochemical indices, inflammatory mediators and the AKI biomarker NGAL; multi-color flow cytometry of peritoneal macrophages (LPS) and intra-renal immune cell subpopulations (CLP) and histology/immunohistochemistry of kidney (CLP). At 72 h post-LPS injections, hUC-MSCs reduced serum inflammatory mediators and peritoneal macrophage M1/M2 ratio. Repeated, but not single, hUC-MSC doses administered at 48 h post-CLP resulted in lower serum concentrations of inflammatory mediators, lower plasma NGAL and reversal of sepsis-associated depletion of intra-renal T cell and myeloid cell subpopulations. Hierarchical clustering analysis of all 48-h serum/plasma analytes demonstrated partial co-clustering of repeated-dose hUC-MSC CLP animals with a Sham group but did not reveal a distinct signature of response to therapy. It was concluded that repeated doses of CD362-selected hUC-MSCs are required to modulate systemic and local immune/inflammatory events in polymicrobial sepsis and SA-AKI. Inter-individual variability and lack of effect of single dose MSC administration in the CLP model are consistent with observations to date from early-phase clinical trials.

Ki-67 as a Prognostic Biomarker in Invasive Breast Cancer.

The advent of molecular medicine has transformed breast cancer management. Breast cancer is now recognised as a heterogenous disease with varied morphology, molecular features, tumour behaviour, and response to therapeutic strategies. These parameters are underpinned by a combination of genomic and immunohistochemical tumour factors, with estrogen receptor (ER) status, progesterone receptor (PgR) status, human epidermal growth factor receptor-2 (HER2) status, Ki-67 proliferation indices, and multigene panels all playing a contributive role in the substratification, prognostication and personalization of treatment modalities for each case. The expression of Ki-67 is strongly linked to tumour cell proliferation and growth and is routinely evaluated as a proliferation marker. This review will discuss the clinical utility, current pitfalls, and promising strategies to augment Ki-67 proliferation indices in future breast oncology.

Gene-eluting stents: adenovirus-mediated delivery of eNOS to the blood vessel wall accelerates re-endothelialization and inhibits restenosis.

Drug-eluting stents for coronary artery disease results in inhibition of smooth muscle cell (SMC) and endothelial cells which may increase the risk of stent thrombosis. In this study, we attempted to enhance re-endothelialization of deployed stents while simultaneously inhibiting intimal hyperplasia by overexpression of endothelial nitric oxide synthase (eNOS) delivery in the vasculature using an adenovirus gene-eluting stent. Re-endothelialization was significantly greater in vessels obtained from normocholesterolemic animals at day 14 (85.34% +/- 7.38 versus 62.66% +/- 10.49; P < 0.05) and day 28 (91.1% +/- 10 versus 63.1% +/- 22; P < 0.05) and hypercholesterolemic animals (96.97% +/- 3.2 versus 28.33% +/- 38.76; P < 0.05) at day 28 with AdeNOS-eluting stents. At day 28, there was a significant increase in the lumen size [AdeNOS 2.73 mm(2) +/- 1.18, AdbetaGal 0.98 mm(2) +/- 0.98, phosphorylcholine (PC) 1.87 mm(2) +/- 1.18; P < 0.05], and a significant reduction in neointimal formation (AdeNOS 2.32 mm(2) +/- 1.13, AdbetaGal 3.73 mm(2) +/- 0.95, PC 3.2 mm(2) +/- 0.94; P < 0.05), and percent restenosis (AdeNOS 45.23 +/- 20.81, AdbetaGal 79.6 +/- 20.31, PC 70.16 +/- 22.2; P < 0.05) in AdeNOS-stented vessels in comparison with controls from hypercholesterolemic animals, assessed by morphometry and quantitative coronary angiography (AdeNOS 15.95% +/- 7.63, AdbetaGal 56.9% +/- 38.6, PC 58 +/- 34.6; P < 0.05). Stent-based delivery of AdeNOS results in enhanced endothelial regeneration and reduction in neointimal formation as compared with controls. This seems to be a promising treatment strategy for preventing in-stent restenosis (ISR) while simultaneously reducing the risk of stent thrombosis.

Practical guide for the comparison of two next-generation sequencing systems for solid tumour analysis in a universal healthcare system.

AIMS: Although there have been excellent reports in the literature of validating next-generation sequencing, comparisons between two systems are not often published due to cost and time. We set out to establish that targetable mutations could be reliably detected with different gene panels and different chemistries using a common bioinformatics pipeline for meaningful comparisons to be made. METHODS: After running selected formalin-fixed, paraffin-embedded samples through QPCR, Sanger sequencing and the 50 gene hotspot v2 panel from Life Technologies to determine standard-of-care variants, we compared the Oncomine panel from Life Technologies performed on a Personal Genome Machine (PGM) and the eight-gene actionable panel from Qiagen performed on a MiSeq platform. We used a common bioinformatics program following the creation of respective VCF files. RESULTS: Both panels were accurate to above 90%, the actionable panel workflow was easier to perform but the lowest effective starting DNA load was obtained on the Oncomine workflow at 4 ng. Such minimal DNA can help with samples where there is limited material such as those for lung cancer molecular studies. We also discuss gene panel content and propose that increasing the gene profile of a panel will not benefit clinical laboratories where standard-of-care testing is all that is required. CONCLUSIONS: Once recognised, it may be cost-effective for such laboratories to begin validation with an appropriate bioinformatics pipeline for targeted multigene hotspot molecular testing.

IHC-based subcellular quantification provides new insights into prognostic relevance of FLIP and procaspase-8 in non-small-cell lung cancer.

In this study, we developed an image analysis algorithm for quantification of two potential apoptotic biomarkers in non-small-cell lung cancer (NSCLC): FLIP and procaspase-8. Immunohistochemical expression of FLIP and procaspase-8 in 184 NSCLC tumors were assessed. Individual patient cores were segmented and classified as tumor and stroma using the Definiens Tissue Studio. Subsequently, chromogenic expression of each biomarker was measured separately in the nucleus and cytoplasm and reported as a quantitative histological score. The software package pROC was applied to define biomarker thresholds. Cox proportional hazards analysis was applied to generate hazard ratios (HRs) and associated 95% CI for survival. High cytoplasmic expression of tumoral (but not stromal) FLIP was associated with a 2.5-fold increased risk of death in lung adenocarcinoma patients, even when adjusted for known confounders (HR 2.47, 95% CI 1.14-5.35). Neither nuclear nor cytoplasmic tumoral procaspase-8 expression was associated with overall survival in lung adenocarcinoma patients; however, there was a significant trend (P for trend=0.03) for patients with adenocarcinomas with both high cytoplasmic FLIP and high cytoplasmic procaspase-8 to have a multiplicative increased risk of death. Notably, high stromal nuclear procaspase-8 expression was associated with a reduced risk of death in lung adenocarcinoma patients (adjusted HR 0.31, 95% CI 0.15-0.66). On further examination, the cells with high nuclear procaspase-8 were found to be of lymphoid origin, suggesting that the better prognosis of patients with tumors with high stromal nuclear procaspase-8 is related to immune infiltration, a known favorable prognostic factor. No significant associations were detected in analysis of lung squamous cell carcinoma patients. Our results suggest that cytoplasmic expression of FLIP in the tumor and nuclear expression of procaspase-8 in the stroma are prognostically relevant in non-small-cell adenocarcinomas but not in squamous cell carcinomas of the lung.

Gene-eluting stents: comparison of adenoviral and adeno- associated viral gene delivery to the blood vessel wall in vivo.

Gene-eluting stents are being evaluated in animals as an alternative approach to inhibiting in-stent restenosis. Adeno-associated virus type 2 (AAV2) and adenovirus are commonly used for gene transfer applications. We tested the hypothesis that these vectors can achieve prolonged and localized gene delivery to the vessel wall, using stents as delivery platforms. AdbetaGal (5 x 10(9) plaque-forming units) and AAV2betaGal (5.3 x 10(9) DNase-resistant particles) were used to coat BiodivYsio stents with matrix HI coating (Abbott Vascular Devices, Galway, Ireland). After balloon injury, external iliac arteries of New Zealand White rabbits were stented. The reverse transcription-polymerase chain reaction was used to assess viral spread. Expression of LacZ was demonstrated with both vectors at five time points (3, 7, 14, 21, and 28 days). In the adenovirus group the median percentage of cells expressing the transgene on day 3 was 2.73%, which increased to a median expression of 7.31% at 28 days (p > 0.05). Expression was localized to medial cells on day 3, but was observed predominantly in neointimal cells on day 28. In the AAV group, day 3 expression was 5.78%, which decreased to 2.12% on day 28 (p = 0.05). No systemic dissemination of virus was seen in any group. Adenovirus- and AAV2-coated stents can be used to deliver genes to the blood vessel wall for up to 28 days.

Characterisation and differentiation of lactobacilli by lectin typing.

Lactobacillus isolates from healthy Estonian and Swedish children were characterised by a lectin typing technique; 56 isolates from six species (L. acidophilus, L. paracasei, L. plantarum, L. fermentum, L. brevis and L. buchneri) were tested. The typing system was based on an agglutination assay with a panel of six commercially available lectins, which were chosen on the basis of their carbohydrate specificities. The isolates were also subjected to proteolytic degradation before lectin typing to decrease auto-agglutination of whole cells in the assay. The 56 isolates were divided into 15 different lectin types by their lectin agglutination patterns. Proteolytic treatment reduced auto-agglutination for the majority of species, apart from L. acidophilus, which remained predominantly auto-agglutinating (eight of nine strains). The system produced stable and reproducible results under standardised culture conditions. Lactobacilli are important bacteria for use as probiotics and this system may supplement current molecular typing techniques and may help in identification of strains that could be useful in this role.

Potential for proteomic profiling of Helicobacter pylori and other Helicobacter spp. using a ProteinChip array.

The Helicobacter genus is associated with a wide spectrum of pathologies in the gastrointestinal tract. However, in contrast to Helicobacter pylori, few data are available regarding proteomic characteristics of enterohepatic helicobacters. Proteomic analysis of this genus has predominantly utilised two-dimensional gel electrophoresis methodology. In the present study we applied an innovative technique using ProteinChip arrays coupled with surface-enhanced laser desorption/ionisation time of flight mass spectroscopy to accurately assess the M(r) of proteins for comparative proteomic profiling. We analysed binding of outer membrane fractions to a weak cation exchange array for strains of H. pylori from culture collections and compared these profiles to fresh clinical isolates. In addition, we analysed profiles from Helicobacter pullorum, Helicobacter bilis and 'Helicobacter sp. flexispira'. The system proved rapid, accurate and reproducible. Distinct specific profiles for all the strains studied were identified. However, strains from culture collections that have undergone numerous subcultures had almost identical profiles. In contrast, profiles from fresh clinical isolates were markedly different. Moreover, certain features of the profiles from the enterohepatic species were conserved.

Through the looking glass: primary epithelioid angiosarcoma in the left atrium, a unique site for a rare malignancy.

Malignant cardiac tumours occurring on the left side are vanishingly rare entities. We describe a case of a 73-year-old male who underwent surgery for a left-sided cardiac tumour following initial presentation with transient ischaemic attacks. In addition to the unusual presentation and subsequent metastatic pattern to the femur, the tumour's pathological diagnosis was that of an epithelioid variant of an angiosarcoma which has not been previously described in this anatomical location.