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Loss of skeletal integrity during ageing and disease is associated with an imbalance in the opposing actions of osteoblasts and osteoclasts1. Here we show that intrinsic ageing of skeletal stem cells (SSCs)2 in mice alters signalling in the bone marrow niche and skews the differentiation of bone and blood lineages, leading to fragile bones that regenerate poorly. Functionally, aged SSCs have a decreased bone- and cartilage-forming potential but produce more stromal lineages that express high levels of pro-inflammatory and pro-resorptive cytokines. Single-cell RNA-sequencing studies link the functional loss to a diminished transcriptomic diversity of SSCs in aged mice, which thereby contributes to the transformation of the bone marrow niche. Exposure to a youthful circulation through heterochronic parabiosis or systemic reconstitution with young haematopoietic stem cells did not reverse the diminished osteochondrogenic activity of aged SSCs, or improve bone mass or skeletal healing parameters in aged mice. Conversely, the aged SSC lineage promoted osteoclastic activity and myeloid skewing by haematopoietic stem and progenitor cells, suggesting that the ageing of SSCs is a driver of haematopoietic ageing. Deficient bone regeneration in aged mice could only be returned to youthful levels by applying a combinatorial treatment of BMP2 and a CSF1 antagonist locally to fractures, which reactivated aged SSCs and simultaneously ablated the inflammatory, pro-osteoclastic milieu. Our findings provide mechanistic insights into the complex, multifactorial mechanisms that underlie skeletal ageing and offer prospects for rejuvenating the aged skeletal system. An analysis of skeletal stem cells in mice reveals that bone ageing occurs at the level of local niches affecting skeletal and haematopoietic lineage output, which may influence systemic aspects of multi-organ physiological ageing.

Supplemental Digital Content is available in the text., SP142 programmed cell death ligand 1 (PD-L1) status predicts response to atezolizumab in triple-negative breast carcinoma (TNBC). Prevalence of VENTANA PD-L1 (SP142) Assay positivity, concordance with the VENTANA PD-L1 (SP263) Assay and Dako PD-L1 IHC 22C3 pharmDx assay, and association with clinicopathologic features were assessed in 447 TNBCs. SP142 PD-L1 intraobserver and interobserver agreement was investigated in a subset of 60 TNBCs, with scores enriched around the 1% cutoff. The effect of a 1-hour training video on pretraining and posttraining scores was ascertained. At a 1% cutoff, 34.2% of tumors were SP142 PD-L1 positive. SP142 PD-L1 positivity was significantly associated with tumor-infiltrating lymphocytes (P

Fragility fractures have a limited capacity to regenerate, and impaired fracture healing is a leading cause of morbidity in the elderly. The recent identification of a highly purified bona fide human skeletal stem cell (hSSC) and its committed downstream progenitor cell populations provides an opportunity for understanding the mechanism of age‐related compromised fracture healing from the stem cell perspective. In this study, we tested whether hSSCs isolated from geriatric fractures demonstrate intrinsic functional defects that drive impaired healing. Using flow cytometry, we analyzed and isolated hSSCs from callus tissue of five different skeletal sites (n = 61) of patients ranging from 13 to 94 years of age for functional and molecular studies. We observed that fracture‐activated amplification of hSSC populations was comparable at all ages. However, functional analysis of isolated stem cells revealed that advanced age significantly correlated with reduced osteochondrogenic potential but was not associated with decreased in vitro clonogenicity. hSSCs derived from women displayed an exacerbated functional decline with age relative to those of aged men. Transcriptomic comparisons revealed downregulation of skeletogenic pathways such as WNT and upregulation of senescence‐related pathways in young versus older hSSCs. Strikingly, loss of Sirtuin1 expression played a major role in hSSC dysfunction but re‐activation by trans‐resveratrol or a small molecule compound restored in vitro differentiation potential. These are the first findings that characterize age‐related defects in purified hSSCs from geriatric fractures. Our results provide a foundation for future investigations into the mechanism and reversibility of skeletal stem cell aging in humans., Human skeletal stem cells (hSSCs) are present and accumulate at fracture sites independent of age. Impaired healing in the elderly might underlie functional defects in hSSCs which are exacerbated in women. Re‐activation of Sirt1 rejuvenates geriatric hSSCs independent of sex.

Conventionally, lymphatic spread is regarded as the principal mechanism by which haematogenous metastasis occurs in colorectal cancer. The aim of this cross sectional study was to determine the relative strengths of direct tumour spread, the presence of lymph node metastasis and histologically demonstrated venous invasion as drivers of haematogenous metastasis diagnosed at the time of resection of colorectal cancer. The data were drawn from a hospital database of consecutive bowel cancer resections between 1995 and 2017 inclusive. The presence of haematogenous metastasis was determined at the time of surgery by imaging or other investigations or operative findings. Where possible, histological confirmation was obtained. Specimen dissection and reporting followed a standardised procedure. Tumour staging was according to the 7th edition of the UICC/AJCC pTNM system. Analysis was by multivariable logistic regression. After exclusions 3133 patients remained, among whom 380 (12.1%) had one or more haematogenous metastases. In bivariate analyses, the frequency of haematogenous metastasis was directly associated with increasing T status (p

Aim The recommended standard of care for patients after resection of Stage III colon cancer is adjuvant 5-fluorouracil based chemotherapy - FOLFOX (fluorouracil, leucovorin with oxaliplatin) - or CAPOX (capecitabine, oxaliplatin). This may be modified in older patients or depending on comorbidity. This has been challenged recently as the apparent benefit of adjuvant chemotherapy may arise from improvements in surgery or preoperative imaging or pathology staging. This study compares recurrence and colon-cancer-specific death between patients who received postoperative adjuvant chemotherapy and those who did not. Method Prospectively recorded data from 363 consecutive patients who had a resection for Stage III colonic adenocarcinoma between 1995 and 2010 inclusive were analysed. Surviving patients were followed for at least 5 years. The suitability of patients for chemotherapy was discussed routinely at multidisciplinary team meetings. The incidence of recurrence and colon-cancer-specific death was evaluated by competing risk methods. Results After adjustment for the competing risk of non-colorectal cancer death, there was no significant difference in recurrence between the 204 patients who received chemotherapy and the 159 who did not [hazard ratio (HR) 0.94, 95% CI 0.66-1.32, P = 0.700) and no significant difference in colon-cancer-specific death (HR 0.73, 95% CI 0.50-1.04, P = 0.084; HR 0.88, 95% CI 0.57-1.36, P = 0.577 after adjustment for relevant covariates). Conclusion These findings question the routine use of chemotherapy after complete mesocolic excision for Stage III colon cancer. Recurrence and cancer-specific death, assessed by competing risk methods, should be the standard outcomes for evaluating the effectiveness of adjuvant chemotherapy after potentially curative resection.

There appears a linear relationship between small increases in running speed and cardiovascular health benefits. Encouraging or coaching recreational runners to increase their running speed to derive these health benefits might be more effective if their joint level kinematic and kinetic strategy was understood. The aim of this investigation was to compare the peak sagittal plane motions, moments, and powers of the hip, knee and ankle at 85%, 100%, 115% and 130% of self-selected running speed. Overground running data were collected in 12 recreational runners (6 women, 6 men) with a full body marker set using a 12-camera Vicon MX system with an AMTI force plate. Kinematics and kinetics were analyzed with Vicon Nexus software. Participants chose to run at 2.6 ± 0.5 m/s (85%); 3.0 ± 0.5 m/s (100%); 3.3 ± 0.5 m/s (115%); and 3.7 ± 0.5 m/s (130%); these four speeds approximately correspond to 6:24-, 5:33-, 5:03-, and 4:30-min kilometer running paces. Running speed had a significant effect (P 0.05) on peak kinematic and kinetic variables of the hips, knees and ankles, with peak sagittal hip moments invariant (P 0.54) and the peak sagittal ankle power generation (P 0.0001) the most highly responsive variable. The timing of the peak sagittal extensor moments and powers at the hip, knee and ankle were distributed across stance in a sequential manner. This study shows that running speed affects lower limb joint kinematics and kinetics and suggests that specific intersegmental kinetic strategies might exist across the narrow range of running speeds.

Aim This study examined pathological quality-of-surgery indicators in laparoscopic and open rectal cancer resection specimens. Methods Retrospective analysis of consecutive, prospectively recorded laparoscopic (LR) or open (OR) resections for rectal cancer. Indicators included integrity of the perirectal fascial envelope, circumferential margin clearance, lymph node yield and distal margin clearance. Results Between January 2007 and December 2013, 168 LR and 189 OR were performed. Univariate analysis showed that the presence of tumor within 1 mm of the circumferential margin was lower in LR (13/168 vs 28/189 cases, P = 0.039) as was a distal margin of clearance of

A 40-year-old woman developed erythematous, raised and pruritic, migratory lesions on a daily basis affecting her whole body for the past 3 months; the appearance of the rash was consistent with urticaria. There was associated angioedema affecting her lips and face. Her medical history was significant for MS, diagnosed 9 years before, manifesting clinically with mobility, speech, and cognitive impairment. Initial treatment was with β-interferon and glatiramer acetate; however, disease relapse prompted switching to alemtuzumab, with 2 standard courses given 12 months apart (60 and 36 mg, respectively, the last dose given 12 months before the onset of rash), and achieved remission with a new baseline Expanded Disability Status Scale of 2.

Aging is characterized by a progressive loss of physiological integrity, leading to impaired function and increased vulnerability to death(1). Despite rapid advances over recent years, many of the molecular and cellular processes which underlie progressive loss of healthy physiology are poorly understood(2). To gain a better insight into these processes we have created a single cell transcriptomic atlas across the life span of Mus musculus which includes data from 23 tissues and organs. We discovered cell-specific changes occurring across multiple cell types and organs, as well as age related changes in the cellular composition of different organs. Using single-cell transcriptomic data we were able to assess cell type specific manifestations of different hallmarks of aging, such as senescence(3), genomic instability(4) and changes in the organism’s immune system(2). This Tabula Muris Senis provides a wealth of new molecular information about how the most significant hallmarks of aging are reflected in a broad range of tissues and cell types.

BACKGROUND: The authors have developed a novel protocol for isolating adipose-derived stem cells from human lipoaspirate. In this study, they compare their new method to a previously published standard protocol. METHODS: Human adipose-derived stem cell isolation was performed using two methods to compare cell yield, cell viability, cell proliferation, and regenerative potential. The new and conventional isolation methods differ in two key areas: die collagenase digestion buffer constituents and the use of an orbital shaker. The osteogenic and adipogenic potential of adipose-derived stem cells isolated using both protocols was assessed in vitro, and gene expression analysis was performed. To assess the ability of the isolated cells to generate bone in vivo, die authors created critical-size calvarial defects in mice, which were treated with adipose-derived stem cells loaded onto hydroxyapatite-coated poly(lactic-co-glycolic acid) scaffolds. To test die ability of the isolated cells to enhance adipogenesis, die cells were added to lipoaspirate and placed beneath the scalp of immunocompromised mice. Fat graft volume retention was subsequently assessed by serial computed tomographic volumetric scanning. RESULTS: The new method resulted in a 10-fold increased yield of adipose-derived stem cells compared with die conventional method. Cells harvested using die new method demonstrated significantly increased cell viability and proliferation in vitro (p < 0.05). New method cells also demonstrated significantly enhanced osteogenic and adipogenic differentiation capacity in vitro (p < 0.05) in comparison with die conventional method cells. Both cell groups demonstrated equivalent osteogenic and adipogenic regenerative potential in mice. CONCLUSIONS: The authors have developed a protocol that maximizes the yield of adipose-derived stem cells derived from lipoaspirate. The new method cells have increased osteogenic and adipogenic potential in vitro and are not inferior to conventional method cells in terms of their ability to generate bone and fat in vivo. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, V.

Aim Complete mesocolic excision (CME) has been advocated as likely to improve the long-term oncological outcome of colon cancer resection, although there is a paucity of long-term results in the literature. The aim of this study was to supplement our previously published results on colon cancer resection based on a standardized technique of precise dissection along anatomical planes with high vascular ligation and to compare our long-term results with those of recent European studies of CME. Method Data were drawn from a prospective hospital registry of consecutive resections for colon cancer between 1996 and 2007, including follow-up to the end of 2012. The principal outcomes from potentially curative resections were 5-year Kaplan–Meier rates of local recurrence, systemic recurrence, overall survival and cancer-specific survival. Secondary outcomes for all resections were postoperative complications, number of lymph nodes retrieved and R0 status. Results For 779 potentially curative resections the local recurrence rate was 2.1% (95% CI 1.3–3.4), the systemic recurrence rate was 10.2% (95% CI 8.1–12.7), the 5-year overall survival rate was 76.2% (95% CI 73.0–79.0) and the cancer-specific survival rate was 89.8% (95% CI 87.3–91.9). For all 905 resections, rates of 14 surgical complications were low and not dissimilar to those in a comparable study. The median lymph node count was 15 (range 0–113). R0 status was confirmed in 883/905 patients (97.6%; 95% CI 96.4–98.5). Conclusion For colon cancer, meticulous dissection along anatomical planes together with high vascular ligation results in few complications, a high R0 rate, low recurrence and high survival.

The pTNM staging system for colorectal cancer (CRC) is not entirely effective in discriminating between potentially curative and non-curative resections because it does not account for local residual tumour in patients with stages I, II or III. This study aimed to evaluate the prognostic importance of histologically verified tumour in any line of resection of the bowel resection specimen (TLR) in relation to pTNM stages and to demonstrate how TLR may be integrated into pTNM staging. Information on patients in the period 1995 to 2010 with complete follow-up to the end of 2015 was extracted from a prospective database of CRC resections. The outcome variables were the competing risks incidence of CRC recurrence and CRC-specific death. After exclusions, 2220 patients remained. In 1930 patients with pTNM stages I-III tumour, recurrence was markedly higher in those with TLR than in those without (HR 6.0, 95% CI 4.2-8.5, p 0.001) and this persisted after adjustment for covariates associated with recurrence. CRC-specific death was markedly higher in the presence of TLR (HR 7.7, CI 5.3-11.2, p 0.001), which persisted after adjustment for relevant covariates. These results justify removing patients with TLR from pTNM stages I to III and placing them in stage IV, thereby allowing the categorisation of all patients with any known residual tumour into three prognostically distinct groups. This study demonstrates how TLR may be integrated into pTNM staging, thus improving the definition of the three stages which are considered potentially curable (I, II and III).

Malignant gastrointestinal neuroectodermal tumour (GNET) is a recently characterised rare and aggressive tumour that typically arises in association with the small intestine of adults. We present a novel case of this entity and expand the spectrum of its reported morphological features. The patient was a 5-year-old female, the youngest reported patient affected by the condition, and presented with extra-abdominal disease. The histopathological features included the presence of a junctional component of the palatal tumour, which mimicked mucosal melanoma, a feature that has not been previously reported in GNET. Whole genome and RNA sequencing was performed that demonstrated the EWSR1-ATF1 translocation characteristic of GNET. Knowledge of this entity and its features, together with careful morphological assessment supplemented by judicious immunohistochemical and molecular studies should enable the correct diagnosis to be established.

Background: cancellation of planned surgery impacts substantially on patients and health systems. This study describes the incidence and reasons for cancellation of inpatient surgery in the UK NHS. Methods: we conducted a prospective observational cohort study over 7 consecutive days in March 2017 in 245 NHS hospitals. Occurrences and reasons for previous surgical cancellations were recorded. Using multilevel logistic regression, we identified patient- and hospital-level factors associated with cancellation due to inadequate bed capacity. Results: we analysed data from 14 936 patients undergoing planned surgery. A total of 1499 patients (10.0%) reported previous cancellation for the same procedure; contemporaneous hospital census data indicated that 13.9% patients attending inpatient operations were cancelled on the day of surgery. Non-clinical reasons, predominantly inadequate bed capacity, accounted for a large proportion of previous cancellations. Independent risk factors for cancellation due to inadequate bed capacity included requirement for postoperative critical care [odds ratio (OR)=2.92; 95% confidence interval (CI), 2.12–4.02; PConclusions: a significant proportion of patients presenting for surgery have experienced a previous cancellation for the same procedure. Cancer surgery is relatively protected, but bed capacity, including postoperative critical care requirements, are significant risk factors for previous cancellations.

Extensive surface profiles of colorectal cancer (CRC) cells and tumor infiltrating lymphocytes (TIL) have been obtained from 45 surgical resection samples. Live cells were captured on an antibody microarray and stained with fluorescently-labeled antibodies. Minimal panels of 11 CRC antigens (CD13, CD24, CD26, CD49d, CD138, CD166, CA-125, CA19-9, EGFR, Galectin-4 and HLA-DR) and 11 T-cell antigens (CD10, CD11b, CD11c, CD25, CD31, CD95, CD151, CD181, Galectin-4, CA19-9, TSP-1) provide signatures for relapse and survival. Hierarchical clustering of profiles from CRC cells and TIL identified groups of patients for survival, systemic relapse and death. The groups from CRC and TIL profiles for systemic relapse showed 79.2% concordance, enabling prediction of relapse after surgery. The results demonstrate communication between CRC cells and TIL.

Diabetes mellitus (DM) is a metabolic disease frequently associated with impaired bone healing. Despite its increasing prevalence worldwide, the molecular etiology of DM-linked skeletal complications remains poorly defined. Using advanced stem cell characterization techniques, we analyzed intrinsic and extrinsic determinants of mouse skeletal stem cell (mSSC) function to identify specific mSSC niche-related abnormalities that could impair skeletal repair in diabetic (Db) mice. We discovered that high serum concentrations of tumor necrosis factor-α directly repressed the expression of Indian hedgehog (Ihh) in mSSCs and in their downstream skeletogenic progenitors in Db mice. When hedgehog signaling was inhibited during fracture repair, injury-induced mSSC expansion was suppressed, resulting in impaired healing. We reversed this deficiency by precise delivery of purified Ihh to the fracture site via a specially formulated, slow-release hydrogel. In the presence of exogenous Ihh, the injury-induced expansion and osteogenic potential of mSSCs were restored, culminating in the rescue of Db bone healing. Our results present a feasible strategy for precise treatment of molecular aberrations in stem and progenitor cell populations to correct skeletal manifestations of systemic disease.

Post-traumatic stress disorder (PTSD) is characterised by a range of debilitating psychological, physical and cognitive symptoms. PTSD has been associated with grey matter atrophy in limbic and frontal cortical brain regions. However, previous studies have reported heterogeneous findings, with grey matter changes observed beyond limbic/frontal areas. Seventy-five adults were recruited from the community, 25 diagnosed with PTSD along with 25 healthy and 25 trauma exposed age and gender matched controls. Participants underwent clinical assessment and magnetic resonance imaging. The data-analyses method Voxel Based Morphometry (VBM) was used to estimate cortical grey matter volumes. When compared to both healthy and trauma exposed controls, PTSD subjects demonstrated decreased grey matter volumes within subcortical brain regions-including the hippocampus and amygdala-along with reductions in the anterior cingulate cortex, frontal medial cortex, middle frontal gyrus, superior frontal gyrus, paracingulate gyrus, and precuneus cortex. Significant negative correlations were found between total CAPS lifetime clinical scores/sub-scores and GM volume of both the PTSD and TC groups. GM volumes of the left rACC and right amygdala showed a significant negative correlation within PTSD diagnosed subjects.

Background Despite an extensive literature suggesting that high microsatellite instability (MSI-H) enhances survival and protects against recurrence after colorectal cancer resection, such effects remain controversial as many studies show only a weak bivariate association or no multivariable association with outcome. This study examined the relationship between MSI status and colorectal cancer outcomes with adjustment for death from other causes as a competing risk. Methods A hospital database of patients following colorectal cancer resection was interrogated for clinical, operative, pathology, adjuvant therapy and follow-up information. MSI-H status was determined by immunohistochemistry for mismatch repair protein deficiency. The cumulative incidence of recurrence and colorectal cancer-specific death was evaluated by competing risks methods. Results Among 1009 patients who had a resection between August 2002 and December 2008, and were followed to at least December 2013, there were 114 (11·3 per cent) with MSI-H (72·8 per cent aged at least 70 years; 63·2 per cent women). After potentially curative resection, with adjustment for non-colorectal cancer death as a competing risk and adjustment for 22 clinical, operative and pathological variables, there was no association between MSI-H and recurrence (hazard ratio (HR) 0·81, 95 per cent c.i. 0·42 to 1·57) or colorectal cancer-specific death (HR 0·73, 0·39 to 1·35) in this patient population. For palliative resections, there was no association between MSI-H and colorectal cancer-specific death (HR 0·65, 0·21 to 2·04). MSI-H was associated with non-colorectal cancer death after both curative (HR 1·55, 1·04 to 2·30) and palliative (HR 3·80, 1·32 to 11·00) resections. Conclusion Microsatellite instability status was not an independent prognostic variable in these patients.

Background: Computerized cognitive behavioral therapy for insomnia (CCBT-I) has a growing evidence base as a stand-alone intervention, but it is less clear what factors may limit its acceptability and feasibility when combined with clinical care. Objective: The purpose of this study was to explore barriers and facilitators to use of an adjunctive CCBT-I program among depressed patients in a psychiatric clinic by using both quantitative and qualitative approaches. Methods: We conducted the qualitative component of the study using face-to-face or telephone interviews with participants who had enrolled in a clinical trial of a CCBT-I program as an adjunctive treatment in a psychiatric clinical setting. In line with the grounded theory approach, we used a semistructured interview guide with new thematic questions being formulated during the transcription and data analysis, as well as being added to the interview schedule. A range of open and closed questions addressing user experience were asked of all study participants who completed the 12-week trial in an online survey. Results: Three themes emerged from the interviews and open questions, consistent with nonadjunctive CCBT-I implementation. Identification with the adjunctive intervention’s target symptom of insomnia and the clinical setting were seen as key reasons to engage initially. Persistence was related to factors to do with the program, its structure, and its content, rather than any nonclinical factors. The survey results showed that only the key active behavioral intervention, sleep restriction, was rated as a major problem by more than 15% of the sample. In this clinical setting, the support of the clinician in completing the unsupported program was highlighted, as was the need for the program and clinical treatment to be coordinated. Conclusions: The use of a normally unsupported CCBT-I program as an adjunctive treatment can be aided by the clinician’s approach. A key behavioral component of the intervention, specific to insomnia treatment, was identified as a major problem for persistence. As such, clinicians need to be aware of when such components are delivered in the program and coordinate their care accordingly, if the use of the program is to be optimized. ClinicalTrial: Australian and New Zealand Clinical Trials Registry ACTRN12612000985886; https://www.anzctr.org.au/Trial/Registration/TrialReview.aspx?id=362875&isReview=true (Archived by WebCite at http://www.webcitation.org/6njjhl42X)

Purpose The aim of this study was to investigate the relationship between tumor budding and other pathology features and overall survival after resection of clinicopathological stage III colon cancer. Methods The number of buds and other histopathological features were assessed in 477 patients who were operated on between 1971 and 2001, with follow-up to December 2006. Overall survival was analyzed using the Kaplan-Meier method and Cox regression. Results The number of buds was dichotomized as low (0 to 8) vs high (>or=9). High budding was more common in men, in high-grade tumors, in the presence of venous invasion, and where the tumor had involved a free serosal surface, but budding was not associated with 8 other clinical and pathological features. The 5-year survival rate for patients with 0 to 8 buds was 51.0% (95% confidence interval, 44.9-55.1), whereas that for patients with 9 or more buds was 33.9% (95% confidence interval, 25.2-42.8). This association, however, disappeared after adjustment for other variables independently associated with survival (hazard ratio, 1.2; 95% confidence interval, 0.94-1.54; P = .139). Conclusion In stage III colon cancer, tumor budding did not provide additional independent prognostic information beyond that given by routine pathology reporting.

Fung C L-S, Chan C, Jankova L, Dent O F, Robertson G, Molloy M, Bokey L, Chapuis P H, Lin B P C & Clarke S J (2010) Histopathology56, 319–330 Clinicopathological correlates and prognostic significance of maspin expression in 450 patients after potentially curative resection of node-positive colonic cancer Aims: The tumour suppressor maspin has been investigated for its association with conventional histopathological features in colorectal cancer and for its potential as an independent predictor of survival and response to adjuvant chemotherapy. The aim of this study was to examine associations between maspin expression, other histopathology and survival in a large consecutive series of patients after potentially curative resection of node-positive colonic adenocarcinoma. Methods and results: Nuclear and cytoplasmic maspin expression in both superficial and deep parts of the tumour were assessed retrospectively by tissue microarray and immunohistochemistry in specimens from 450 patients whose other histopathology had been recorded in a prospective hospital registry of large bowel cancer resections from 1971 to 2001 with a minimum follow-up of 5 years. Among 13 clinicopathological features examined, the only associations that persisted across all four maspin assessments were stronger expression in right- than in left-sided tumours (P = 0.001–0.011) and stronger expression in high-grade tumours (P

Aims To investigate the role of DNA repair proteins and their prognostic significance in non-small-cell lung cancer (NSCLC). Methods and results A retrospective analysis of 108 cases of stage I–II NSCLC was undertaken. Immunohistochemical expression of DNA repair proteins MLH1, MSH2 and MGMT was assessed using tissue microarrays of paraffin-embedded samples of invasive carcinoma and precursor lesions. Results were analysed in relation to clinicopathological parameters and patient survival. Reduced expression of MLH1 was found in 58.5% of tumours and occurred less frequently in poorly differentiated tumours (P = 0.044) and large cell carcinomas (P = 0.004). MSH2 and MGMT expression was reduced in 18.1% and 77.8% of cases, respectively. There was an inverse relationship between MLH1 and MSH2 expression (P = 0.012). Normal expression of MLH1, MSH2 and MGMT was found in all cases of squamous metaplasia and squamous dysplasia. Only a single case of carcinoma in situ (12.5%) showed reduced MLH1, none showed reduced MSH2 and 25% showed reduced MGMT. Survival analyses showed no prognostic significance based on expression of MLH1 (P = 0.92), MSH2 (P = 0.78) or MGMT (P = 0.57). Conclusions Reduction in expression of DNA repair proteins MLH1, MSH2 and MGMT is relatively common in NSCLC, appears to be a late event in the development of invasive malignancy and does not influence survival in this patient cohort. Cooper W A, Kohonen-Corish M R J, Chan C, Kwun S Y, McCaughan B, Kennedy C, Sutherland R L & Lee C-S (2008) Histopathology52, 613–622 Prognostic significance of DNA repair proteins MLH1, MSH2 and MGMT expression in non-small-cell lung cancer and precursor lesions

Background: The aim of this study was to determine the extent to which pathology reporting of colorectal cancers notified to the New South Wales Central Cancer Registry during 2000 conformed to guidelines promulgated by the National Health and Medical Research Council. Methods: De-identified reports for 2233 resected specimens of primary invasive colorectal carcinoma were coded according to a standardized system to compile information on 28 clinical and pathology features. An overall score for each report was calculated by computing the number out of 13 essential features specified in the guidelines for which data had been recorded explicitly and unambiguously in the report. Results: The overall score ranged from 3 to 13 features with a mean of 9. No more than 7 features were reported explicitly in just less than one quarter of the reports and no more than 10 in three quarters. There were only 110 reports (4.9%) that included all features. Information on direct spread and nodal metastasis was well reported; resection margins less so. Many reports lacked information on metastases beyond the operative field, the involvement of deep or circumferential resection margins and tumour stage. Conclusion: In some respects pathology reports of resected colorectal cancer specimens displayed a high level of completeness. Some important features, however, were poorly described. Reporting could be improved if surgeons were to use a standardized form to convey clinical information to the pathologist and if pathologists were to report in a structured or synoptic format, explicitly recording the presence or absence of each feature in a standard list.

Objective To determine whether the presence of tumour at a free serosal surface was independently associated with pelvic recurrence or survival in patients who had a resection for clinicopathological stage B or stage C rectal cancer and who had not received adjuvant therapy. Method Data were drawn from a comprehensive, prospective hospital registry of all resections for rectal cancer from January 1971 to December 1998 with follow up to December 2003. Statistical analysis employed the χ2 test or Fisher's exact probability, Kaplan–Meier estimation and proportional hazards regression, with a significance level of ≤0.05 and 95% confidence intervals (CI). Results In 665 patients with stages B or C tumour, 35 (5.3%; CI 3.7–7.2%) had tumour at a free serosal surface. These comprised 6/332 (1.8%; CI 0.8–3.7%) patients with stage B tumour and 29/333 (8.7%; CI 6.1–12.2%) with stage C tumour. After adjustment for other relevant variables, involvement of a free serosal surface was significantly associated with pelvic recurrence [hazard ratio (HR) 2.7; CI 1.3–5.5] and diminished survival (HR 1.6; CI 1.1–2.4) but not with systemic (only) recurrence. Conclusion This study has confirmed that direct tumour spread to a free serosal surface independently predicts pelvic recurrence and diminished survival after resection of clinicopathological stage B and C rectal cancer. This feature should always be sought by the pathologist and reported when present, and noted by the surgeon and oncologist. Serosal involvement should be evaluated further for its utility in selecting patients for adjuvant therapy.

© 2015, Society of General Internal Medicine. Background: Reducing patient cost-sharing and engaging patients in disease management activities have been shown to increase uptake of evidence-based care. Objective: To evaluate the effect of employer purchase of a disease-specific plan with reduced cost-sharing and disease management (the Diabetes Health Plan/DHP) on medication adherence among eligible employees and dependents. Design: Employer-level “intent to treat” cohort study, including data from eligible employees and their dependents with diabetes, regardless of whether they were enrolled in the DHP. Setting: Employers that contracted with a large national health plan administrator in 2009, 2010, and/or 2011. Participants: Ten employers that purchased the DHP and 191 employers that did not (controls). Inverse probability weighting (IPW) estimation was used to adjust for inter-group differences. Intervention: The DHP includes free or low-cost medications and physician visits. Enrollment strategies and specific benefit designs are determined by the employer and vary in practice. DHP participants are notified up front that they must engage in their own health care (e.g., receiving diabetes-related screening) in order to remain enrolled. Main Outcome Measure: Mean employee adherence to metformin, statins, and ACE/ARBs at the employer level at one year post-DHP implementation, as measured by the proportion of days covered (PDC). Results: Baseline adherence to the three medications was similar across DHP and control employers, ranging from 64 to 69 %. In the first year after DHP implementation, predicted employer-level adherence for metformin (+4.9 percentage points, p = 0.017), statins (+4.8, p = 0.019), and ACE/ARBs (+4.4, p = 0.02) was higher with DHP purchase. Limitations: Non-randomized, observational study. Conclusions: The Diabetes Health Plan, an innovative health plan that combines reduced cost-sharing and disease management with an up-front requirement of enrollee participation in his or her own health care, is associated with a modest improvement in medication adherence at 12 months.

ObjectivesThe purpose of this study was to evaluate whether radiation exposure from cardiac computed tomographic angiography (CTA) is associated with deoxyribonucleic acid (DNA) damage and whether damage leads to programmed cell death and activation of genes involved in apoptosis and DNA repair.BackgroundExposure to radiation from medical imaging has become a public health concern, but whether it causes significant cell damage remains unclear.MethodsWe conducted a prospective cohort study in 67 patients undergoing cardiac CTA between January 2012 and December 2013 in 2 U.S. medical centers. Median blood radiation exposure was estimated using phantom dosimetry. Biomarkers of DNA damage and apoptosis were measured by flow cytometry, whole genome sequencing, and single cell polymerase chain reaction.ResultsThe median dose length product was 1,535.3 mGy·cm (969.7 to 2,674.0 mGy·cm). The median radiation dose to the blood was 29.8 mSv (18.8 to 48.8 mSv). Median DNA damage increased 3.39% (1.29% to 8.04%, p < 0.0001) and median apoptosis increased 3.1-fold (interquartile range [IQR]: 1.4- to 5.1-fold, p < 0.0001) post-radiation. Whole genome sequencing revealed changes in the expression of 39 transcription factors involved in the regulation of apoptosis, cell cycle, and DNA repair. Genes involved in mediating apoptosis and DNA repair were significantly changed post-radiation, including DDB2 (1.9-fold [IQR: 1.5- to 3.0-fold], p < 0.001), XRCC4 (3.0-fold [IQR: 1.1- to 5.4-fold], p = 0.005), and BAX (1.6-fold [IQR: 0.9- to 2.6-fold], p < 0.001). Exposure to radiation was associated with DNA damage (odds ratio [OR]: 1.8 [1.2 to 2.6], p = 0.003). DNA damage was associated with apoptosis (OR: 1.9 [1.2 to 5.1], p < 0.0001) and gene activation (OR: 2.8 [1.2 to 6.2], p = 0.002).ConclusionsPatients exposed to >7.5 mSv of radiation from cardiac CTA had evidence of DNA damage, which was associated with programmed cell death and activation of genes involved in apoptosis and DNA repair.

Purpose The significance of low microsatellite instability (MSI-L) in colorectal cancer is poorly understood. No clear biologic distinction has been found between MSI-L and microsatellite stable (MSS) colorectal cancer, and these two phenotypes are usually combined when analyzed against the well-defined high MSI (MSI-H) phenotype. Evidence is emerging that an O6-methylguanine DNA methyltransferase (MGMT) gene defect is associated with MSI-L. Therefore, to further define this phenotype, we undertook a detailed analysis of the prognostic significance of MSI-L and loss of MGMT expression in colon cancer. Patients and Methods The study cohort was 183 patients with clinicopathologic stage C colon cancer who had not received adjuvant therapy. We analyzed MSI status, MGMT, and mismatch repair protein expression, as well as MGMT and p16 promoter hypermethylation. Results We showed that MSI-L defines a group of patients with poorer survival (P = .026) than MSS patients, and that MSI-L was an independent prognostic indicator (P = .005) in stage C colon cancer. Loss of MGMT protein expression was associated with the MSI-L phenotype but was not a prognostic factor for overall survival in colon cancer. p16 methylation was significantly less frequent in MSI-L than in MSI-H and MSS tumors and was not associated with survival. Conclusion MSI-L characterizes a distinct subgroup of stage C colon cancer patients, including the MSI-L subset of proximal colon cancer, who have a poorer outcome. Neither the MGMT defect nor p16 methylation are likely to contribute to the worse prognosis of the MSI-L phenotype.

The purpose of this study is to demonstrate safety and effectiveness of the S-Stent in de novo coronary lesions treated with conventional percutaneous coronary balloon angioplasty. Between January 2000 and June 2001, 120 patients were prospectively enrolled at four study centers. Patients were treated with coronary stenting in a total of 137 lesions. Procedural success was achieved in 100% of 137 attempted lesions. Clinical success was 99.8%. In-hospital mortality was 0.8%; myocardial infarction occurred in 0.8% and stent thrombosis in 0.8%. After stent implantation, the minimal lumen diameter increased from 0.92 ± 0.43 to 2.74 ± 0.36 mm (P < 0.0001) and the percent diameter stenosis decreased from 68.0 ± 16.2 to 4.5 ± 12.0 (P < 0.0001). At 6-month follow-up, the percent diameter stenosis was 33.5 ± 21.3 and the angiographic restenosis rate was 16.5%. Target lesion revascularization was required in 12 patients (10.1%). We conclude that the use of S-Stent for coronary intervention resulted in a high procedural success rate and low angiographic restenosis at 6 months after implantation. Catheter Cardiovasc Interv 2004;62:439–444. © 2004 Wiley-Liss, Inc.

This study should clarify whether the gold-coated NIROYAL stent is equivalent to the stainless steel NIR stent. Patients were randomized to either NIR stent (n = 298) or a NIROYAL stent (n = 305). The primary endpoint was the minimum lumen diameter of the target lesion at 6 months postprocedure. Secondary endpoints focused on clinical events. At 30 days, adverse events were similar in both groups. At 6 months, the minimal lumen diameter was 1.83/1.64 mm (P < 0.001; 95% CI = 0.08-0.30) and the angiographic restenosis rate was 20.6%/37.7% (P < 0.001; 95% CI = -24.7 to -9.3) for NIR/NIROYAL. The 6-month MACE rates were NIR 7.4% and NIROYAL 10.5% (95% CI = -7.7 to 1.4). Compared to stainless steel stent, the NIROYAL stent demonstrated a smaller minimal lumen diameter, a higher late loss (i.e., higher neointimal hyperplasia in spite of a significantly better initial gain), with higher restenosis and similar MACE rates at 6 months.

We conducted a randomized feasibility pilot study comparing combined fibrinolysis with immediate rescue angioplasty vs. primary angioplasty with adjunctive abciximab in patients with acute myocardial infarction (AMI). Seventy patients with ST segment elevation AMI of ≤ 6 hr were randomized to either 50 mg of alteplase and abciximab (n = 34) or primary angioplasty with adjunctive abciximab (n = 36). Coronary angiography was performed at 60 min in the combined lytic group and TIMI 3 flow was present in 65% of patients as compared to 25% (P = 0.001) in the primary angioplasty group prior to intervention. Treatment success, defined as TIMI 3 flow, was achieved in 83% of patients in the primary angioplasty group (P = 0.075 compared to 65% in combined lytic group before rescue angioplasty). There was no difference in overall treatment success between primary angioplasty and combined lytic group with rescue angioplasty (83% vs. 94%; P = NS). Major adverse cardiac events at 1 month were not significant (15% vs. 11%; P = NS), but there was a trend toward more events in the combined lytic group at 6 months (32% vs. 14%; P = 0.066), particularly in target vessel revascularization. In this feasibility pilot study, high rate of TIMI 3 flow was attained in patients with AMI with both combined fibrinolysis and primary angioplasty with adjunctive abciximab. A larger randomized trial is currently ongoing to compare these two strategies. Cathet Cardiovasc Intervent 2004;62:445–452. © 2004 Wiley-Liss, Inc.

The purpose of this study was to determine the impact of these 2 reperfusion strategies (reduced-dose alteplase plus abciximab or direct angioplasty plus abciximab) on fibrinolytic and thrombin generation activities. The effect of reduced-dose alteplase plus abciximab and direct angioplasty plus abciximab on hemostatic factors is unknown. Of 70 patients with acute myocardial infarction ofor = 6 hours, 34 were randomized to reduced-dose alteplase (35 to 50 mg in 1 hour) and 36 to direct angioplasty. A standard bolus and infusion dose of abciximab was administered to all patients. Blood specimens were collected at baseline, and at 1, 4, 12, and 24 hours. The following parameters were assayed: fibrinogen, plasminogen and antiplasmin activities, tissue plasminogen activator antigen, D-dimer, prothrombin fragments F1 + 2, and thrombin/antithrombin III complexes. Among patients treated with reduced-dose alteplase plus abciximab, the fibrinogen level decreased by 28.4% in the first hour (11.7 +/- 3.4 vs 7.8 +/- 2.5 micromol/L, p0.001). Correspondingly, plasminogen and antiplasmin activities decreased by 43.8% (p0.001) and 59.1% (p0.001), respectively. Prothrombin fragments F1 + 2 increased from 2.2 +/- 1.7 to 4.2 +/- 1.6 nmol/L (1 hour) (p0.001) and thrombin/antithrombin III increased from 16.3 +/- 15.0 to 33.5 +/- 19.9 microg/L (1 hour) (p0.001). Conversely, in the direct angioplasty group, there was a marginal elevation in fibrinogen level at 1 hour (10.2 +/- 2.4 vs 10.6 +/- 2.0 micromol/L, p = 0.064) despite a significant reduction in plasminogen and an increase in tissue plasminogen activator levels. There was no significant change in prothrombin fragments F1 + 2 and thrombin/antithrombin III levels. Thus, there was considerable fibrinolytic activity with reduced-dose alteplase plus abciximab; thrombin generation was not prevented. Among patients treated with direct angioplasty, there was some endogenous fibrinolytic activity, but there was no significant thrombin generation.

Aims The diagnosis of intraductal papillary lesions of the breast on core biopsy remains challenging in pathology, with most patients requiring formal surgical excision for a definitive diagnosis. The aim of this study was to determine whether a representative panel of proliferative cell cycle immunohistochemical markers (cyclin A2, cyclin B1 and cyclin D1) could improve the specificity of pathological diagnosis of these lesions. Methods A series of 68 surgically excised intraductal papillary lesion cases were retrospectively selected, and immunohistochemistry for cyclin A2, cyclin B1 and cyclin D1 was performed. Results Cyclin B1 (OR 1.80, 95% CI 1.01 to 3.2, p=0.046) and cyclin D1 (OR 1.13, 95% CI 1.05 to 1.22, p=0.002) expression was independently associated with a diagnosis of malignancy in papillary lesions, although expression was frequently heterogeneous and only focal. Cyclin A2 expression (OR 0.76, 95% CI 0.41 to 1.4, p=0.38) was not associated with a malignant diagnosis in multivariable logistic regression models. All three cyclins displayed high sensitivity (80%–95%) for a diagnosis of malignancy, although cyclin B1 showed a superior specificity of 72.7% compared with the low specificity of cyclins A2 and D1. Conclusions Our study has identified for the first time that the expression of key cell cycle markers differs between benign and malignant papillary breast lesions and identified changes to the mitotic marker, cyclin B1, as particularly significant. However, given the low level and heterogeneous nature of expression of these markers, there remains a significant risk of undersampling in core biopsies and thus they are unlikely to be useful in routine clinical practice.

Extramural venous invasion is a known independent predictor of poor prognosis after resection of colorectal adenocarcinoma, but the prognostic value of mural venous invasion alone and the association between venous invasion and prognosis within tumor stages has received little research attention.This study aimed to determine whether associations between mural and extramural venous invasion and outcome differ among tumor stages after adjustment for other factors known to influence prognosis.This study is a retrospective analysis of prospectively collected data.Data were drawn from a registry of 3040 consecutive patients undergoing resection between 1980 and 2005 under the care of specialist surgeons in a tertiary referral public hospital and an affiliated private hospital. A standardized protocol was used for the pathological assessment of specimens.The primary outcomes measured were overall survival, cancer-specific survival, and recurrence.There was no significant association between venous invasion and survival in stages A (n = 544) or B (n = 1078). In stage C (n = 899), overall survival time was significantly shorter in patients with mural invasion alone or extramural invasion (both p0.001) than in those without invasion, and this persisted after adjustment for other prognostic variables. Equivalent bivariate associations were found in stage D, but only the effect of extramural invasion persisted after adjustment.Our findings arise from the experience of a single surgical group and may not be generalizable to other settings. Only hematoxylin and eosin staining was used.The association between venous invasion and prognosis was stage specific. Both mural venous invasion alone and extramural venous invasion independently predicted overall survival in patients with stage C tumors, but not in patients with stages A, B, or D tumors. Although mural invasion alone was rare, the separate reporting of both mural and extramural invasion in patients with stage C tumor is informative and desirable.

Adipose tissue represents an abundant and easily accessible source of multipotent cells that may serve as an excellent building block for tissue engineering. However, adipose-derived stromal cells (ASCs) are a heterogeneous group and subpopulations may be identified with enhanced osteogenic potential.Human ASC subpopulations were prospectively isolated based on expression of bone morphogenetic protein receptor type-IB (BMPR-IB). Unsorted, BMPR-IB(+), and BMPR-IB(-) cells were analyzed for their osteogenic capacity through histological staining and gene expression. To evaluate their in vivo osteogenic potential, critical-sized calvarial defects were created in immunocompromised mice and treated with unsorted, BMPR-IB(+), or BMPR-IB(-) cells. Healing was assessed using microcomputed tomography and pentachrome staining of specimens at 8 weeks.Increased osteogenic differentiation was noted in the BMPR-IB(+) subpopulation, as demonstrated by alkaline phosphatase staining at day 7 and extracellular matrix mineralization with Alizarin red staining at day 14. This was also associated with increased expression for osteocalcin, a late marker of osteogenesis. Radiographic analysis demonstrated significantly enhanced healing of critical-sized calvarial defects treated with BMPR-IB(+) ASCs compared with unsorted or BMPR-IB(-) cells. This was confirmed through pentachrome staining, which revealed more robust bone regeneration in the BMPR-IB(+) group.BMPR-IB(+) human ASCs have an enhanced ability to form bone both in vitro and in vivo. These data suggest that positive selection for BMPR-IB(+) and manipulation of the BMP pathway in these cells may yield a highly osteogenic subpopulation of cells for bone tissue engineering.

Cancer sufferers are often found to use herbal products along with targeted therapy although not much information (whether beneficial or harmful) is available about the effects of such combinations. In this study, we investigated synergism from the combination of platinum drugs and a number of tumour-active phytochemicals including curcumin, epigallocatechin-3-gallate, thymoquinone, genistein, resveratrol, betulinic acid and ursolic acid in three human ovarian cancer cell lines A2780, A2780(cisR) and A2780(ZD0473R), as a function of concentration and the sequence of administration. Both the dose-effect curves and combination indices show that the binary combinations of platinum drugs with the phytochemicals exert concentration- and sequence-dependent synergism in the cell lines. Generally the degree of synergism is found to be greater in sequenced administration such as 0/2 h, 2/0 h, 0/4 h and 4/0 h than the bolus. The variation in the nature of the combined drug action from being highly synergistic to antagonistic with the change in sequence of administration clearly indicates that the action of one drug modulates that of the other (towards the induction or inhibition of apoptosis). We have also used sequenced combinations of platinum drugs and bortezomib (a proteasome inhibitor that prevents cisplatin-induced proteasomal degration of copper transporter CTR1) to enhance cellular platinum accumulation and the level of platinum-DNA binding especially in the resistant human ovarian tumour models. Proteomic studies to identify the key proteins associated with platinum resistance are ongoing. We have identified 59 proteins associated with platinum resistance in ovarian tumor models.

Background The preoperative ratio of neutrophils to lymphocytes (NLR) has been proposed as a marker of poor outcome in patients having a resection for colorectal cancer (CRC). This study investigated the association between NLR and overall survival, cancer-specific survival and recurrent cancer in patients who had a potentially curative resection for node-positive CRC. Methods Data on 322 patients were drawn from a prospectively recorded registry operated on between 1999 and 2007. Analyses of survival involved the Kaplan-Meier method, Cox regression and competing risks Cox regression. Results Increasing NLR as a continuous variable was independently though weakly associated with diminishing overall survival after adjustment for other prognostic variables (HR 1.06, 95% CI 1.01-1.11, p = 0.013). Receiver operating characteristic analysis to dichotomize NLR as a predictor of overall survival yielded relatively poor sensitivity (55%), specificity (66%) and positive predictive value (56%, CI 47%-64%). Competing risks regression also showed that NLR was not independently associated with recurrence at any site (HR 1.04, CI 0.97-1.11, p = 0.241) or CRC-specific mortality (HR 1.02, CI 0.92-1.12, p = 0.782) but was associated with non-CRC mortality (HR 1.09, CI 1.03-1.15, p = 0.004). Conclusion In patients with stage C tumor the weak link between NLR and overall mortality was not specific to CRC but apparently arose because patients with an elevated inflammatory status preoperatively were likely to progress to earlier death but not necessarily because of their cancer.

Chemopreventative phytochemicals having antitumour and antioxidant properties can overcome problems of chemoresistance and nonspecific toxicity towards normal cells that are associated with platinum-based chemotherapy against cancer. These agents exert their effects by bringing into play numerous cellular proteins that in turn affect multiple steps in pathways leading to tumourigenesis. In this study, combinations of two cytotoxic phytochemicals anethole and curcumin were applied in binary combination with platinum drugs cisplatin and oxaliplatin to three epithelial ovarian cancer cell lines: A2780 (parent), A2780(cisR) (cisplatin-resistant) and A2780(ZD0473R) (ZD0473-resistant). Cell viability was quantified using the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) reduction assay and the combined drug action was analyzed based on the equations derived by Chou and Talalay (1984). Greatest synergism was observed when the phytochemical was added first followed by the platinum drug 2 h later and additiveness to antagonism in combined drug action was observed when the two compounds were administered as a bolus. If confirmed in vivo, the appropriate sequenced combinations of platinum with the phytochemicals may provide a means of overcoming drug resistance.

In this study, synergism in activity from the sequenced combinations of three trans-palladiums (denoted as TH5, TH6 and TH7) with green tea polyphenol (-)-epigallocatechin-3-gallate (EGCG), as well as that with cisplatin, was investigated in a number of human ovarian tumour models as a function of sequence of administration. Cellular accumulation of platinum and palladium, and the levels of platinum-DNA and palladium-DNA binding were also determined for the 0/4 h and 0/0 h sequences of administration. The results of the study show that co-administration of cisplatin with EGCG (0/0 h) produces weak synergism in both cisplatin-sensitive (A2780) and cisplatin-resistant (A2780(cisR)) cell lines whereas (0/4 h) administration produces pronounced synergism in both. In contrast, bolus administration of EGCG with TH5, TH6 and TH7 produces marked antagonism except that with TH5, in the A2780(cisR) cell line, where a mild synergism is observed. In the case of TH5, TH6 and TH7, administration of drugs with a time gap (0/4 h or 4/0 h combinations) produces sequence-dependent synergism in both A2780 and A2780(cisR) cell lines, whereas in the case of cisplatin, marked antagonism is observed with the 4/0 h sequence of administration in the A2780 cell line. Whereas the highly synergistic 0/4 h sequence of combination of cisplatin with EGCG is found to be associated with pronounced cellular accumulation of platinum and a high level of platinum-DNA binding, no such clear trend can be seen for any of the combinations of TH5, TH6 and TH7 with EGCG. The results of the present study provide support to the idea that sequenced combinations of platinum drugs and tumour-active palladium compounds with selected phytochemicals such as EGCG may provide a means of overcoming drug resistance.

The present study deals with the synthesis, characterization, and activity against human ovarian cancer cell lines A2780, A2780(cisR), A2780(ZD0473R), and SKOV-3 of three trans-planaramine-palladium(II) complexes of the form trans-PdL(2)Cl(2), coded as EH1, EH3, and EH4, for which L = 2-methylpyridine, imidazole, and 1,2-α-imidazopyridine, respectively. The cellular accumulation of palladium, palladium-DNA binding levels, and the nature of interactions of the compounds with salmon sperm and pBR322 plasmid DNA were also determined. All three compounds were found to be less active than cisplatin, but unlike cisplatin they were found to be equally or more active against the resistant cell lines A2780(cisR) and A2780(ZD0473R) than against the parent cell line A2780. Among the three palladium complexes, EH4 (which has the bulkiest carrier ligand) was found to be most active, in line with the highest cellular accumulation of palladium and highest level of palladium-DNA binding resulting from the compound. EH4 was also found to cause the greatest conformational change to pBR322 plasmid DNA. The results of this study illustrate structure-activity relationships; in particular, they support the idea that the decreased reactivity of trans-palladium complexes through the introduction of bulky ligands can make them more active against tumors.

Profound hypercalcemia is usually due to underlying malignancy.We describe a case of granulomatous myositis presenting with extreme hypercalcemia of 20.1 mg/dl and generalized weakness that did not resolve despite rapid correction of serum calcium. The disease process was unmasked by cholecalciferol supplementation. Initial search for a malignant process yielded no diagnosis, but an elevated 1,25-dihydroxyvitamin D level, in the setting of a suppressed PTH and undetectable PTHrP, pointed to the presence of excessive 1α-hydroxylase activity.Biopsy of the vastus lateralis muscle showed extensive granulomatous myositis. Immunohistochemical staining for 1α-hydroxylase was localized to the multinucleated giant cells and histiocytes. Musculoskeletal magnetic resonance imaging showed involvement of proximal muscle groups of both thighs and upper limbs.Measurement of vitamin D metabolites is pivotal in diagnosing 1,25-dihydroxyvitamin D-mediated hypercalcemia. Granulomatous disease can occasionally cause profound hypercalcemia and needs to be considered in the differential diagnosis. 1,25-Dihydroxyvitamin D-mediated hypercalcemia is responsive to glucocorticoid therapy.

S100A4, a multifunctional protein, has been linked to the invasive growth and metastases of several human cancers. This study investigated the association between S100A4 and overall survival and other clinicopathological features in patients with stage C colonic cancer.Clinical and pathological data were obtained from a prospective hospital registry of 409 patients who had a resection for stage C colonic cancer. Tissue microarrays for immunohistochemistry were constructed from archived tissue. S100A4 staining intensity and percentage of stained cells were assessed in nuclei and cytoplasm for both the central part of the tumour and at the advancing front. Overall survival was analysed by the Kaplan-Meier method and Cox regression.Only a high percentage of cells with S100A4 cytoplasmic staining in frontal tissue was associated with poor survival (hazard ratio, 1.6; 95 % CI 1.1-2.2; p = 0.008) after adjustment for other prognostic variables. There was no association between frontal cytoplasmic S100A4 expression and any of 13 other clinicopathological variables.High expression of S100A4 in cytoplasm at the advancing front of stage C colonic tumours indicates a poor prognosis. Whether S100A4 can predict response to adjuvant chemotherapy remains to be investigated in a randomised clinical trial.

This study investigated the association between glutathione S-transferase Pi (GST Pi) expression, histopathology and overall survival in 468 patients after resection of stage C colonic adenocarcinoma.Data were drawn from a prospective hospital registry of consecutive bowel cancer resections with a minimum follow-up of 5 years. Nuclear and cytoplasmic GST Pi expression, assessed by both intensity of staining and percentage of stained cells at both the central part of the tumour and the invasive tumour front, were evaluated retrospectively by tissue microarray immunohistochemistry on archival specimens. The most effective measure of GST Pi expression was the percentage of immunostained nuclei in central tumour tissue, where40% stained was associated significantly with high grade, invasion beyond the muscularis propria, involvement of a free serosal surface or apical node, and invasion into an adjacent organ or structure. After adjustment of other predictors, GST Pi expression remained independently prognostic for reduced overall survival (hazard ratio 1.4, P = 0.002).In patients with clinicopathological stage C colonic cancer, GST Pi expression is associated with features of tumour aggressiveness and with reduced overall survival. Further appropriately designed studies should aim to discover whether GST Pi can predict response to adjuvant chemotherapy.

The development of drug resistance remains one of the major hurdles in cancer chemotherapy, particularly so for ovarian cancer. Combination of drugs acting synergistically in combination can offer a means of overcoming drug resistance. In this study, two tumour-active phytochemicals, quercetin and thymoquinone, were combined with two platinum drugs, cisplatin and oxaliplatin, with the aim of providing a means of overcoming drug resistance. Two human epithelial ovarian cancer cell lines, A2780 and its cisplatin-resistant form (A2780(cisR)) were treated with binary combinations of cisplatin and oxaliplatin with quercetin and thymoquinone using three sequences of administration. Cell viability was quantified using the (MTT) reduction assay. The combined drug action was analysed based on the equations derived by Chou and Talalay (1984). Greatest synergism was observed when the phytochemical was added first followed by platinum drug 2 h later and the least synergism (often additive to antagonistic) was observed when the two compounds were administered as a bolus. It is suggested that the addition of the phytochemical 2 h before platinum drug may sensitize cancer cells to platinum action, thus offering a means of overcoming drug resistance. The results may be highly significant clinically if found to be confirmed in vivo.

The influence of balloon aortic valvuloplasty (BAV) on aortic regurgitation (AR) in patients with severe aortic stenosis associated with greater than or equal to grade II AR was studied by supraaortic angiogram before and after BAV. The results of 50 patients aged 72 +/- 12 years with significant AR before BAV (group A) were compared to 297 patients (mean age 76 +/- 10 years) with no or mild AR (group B). In group A, the patients had a higher left ventricular end diastolic volume (96 +/- 19 mL/m 2 vs 81 +/- 32 mL/m 2, P less than 0.01) and left ventricular end diastolic pressure (23 +/- 9 mmHg vs 19 +/- 9 mmHg, P less than 0.01). The aortic valve area was similar in both groups. Following BAV, the improvement in aortic valve area and hemodynamics were similar in both groups. In group A, AR remained unchanged in 31 patients (62%), increased by 1 grade in 13 patients (26%), and decreased by 1 grade in 6 patients (12%). In group B, AR increased by greater than 1 grade in 34 patients (11%) and greater than 2 grades in 4 patients (1.3%) post-BAV. Two patients in group B underwent emergency aortic valve replacement following BAV because of severe acute AR. In conclusion, when it is indicated, BAV can be performed with similar risk in patients with significant AR.

Fascin, an actin-bundling protein, is expressed in many neoplasms including colorectal cancer. It is considered to be a mediator of tumor cell invasion and an indicator of aggressive phenotype; however, there are few reports on the association between fascin and prognosis in colorectal cancer. The aims of this study were to: (a) investigate the expression of fascin in the central part of the tumor and at the invasive front in patients who had a potentially curative resection for node-positive colonic carcinoma; (b) examine the method of scoring fascin expression; and (c) investigate the association between fascin expression and overall survival and other clinicopathologic features. Fascin expression was assessed by immunostaining ofmicroarrays from archived tissue of 470 patients who were followed for a minimum of 5 years after resection. Other clinicopathologic data had been recorded prospectively according to a standardized protocol. Analysis of overall survival was by the Kaplan-Meier method and Cox regression. For both central tumor tissue and the invasive front, it was found that the percentage of stained cells was a sufficient measure of fascin expression in relation to survival, with staining intensity providing no significant additional information. At both levels, there was a significant independent association between high fascin expression and diminished survival, although this association was much stronger in the central region (adjusted hazard ratio 1.6, P < 0.001) than at the invasive front (adjusted hazard ratio 1.1, P = 0.044). Fascin expression predicted overall survival but did not displace other routinely collected clinicopathologic predictors.

Objectives. The goal of the study was to evaluate the progressive increase in ischemic threshold with multiple sequential transient coronary occlusions and to assess the role of the collateral circulation in adaptation to ischemia. Background. It has been observed that the duration of balloon inflations during coronary angioplasty can be gradually prolonged during subsequent dilations with a reduction in patient symptoms and diminished ischemic electrocardiographic (ECG) changes. Although the mechanism has not been fully explained, recruitment of coronary collateral circulation induced by repeated coronary occlusion has been reported. The stimuli for recruitment and the natural history of coronary collateral circulation are not understood. Methods. Seventeen patients with isolated stenosis of the left anterior descending coronary artery and a normal left ventricle were enrolled. Angioplasty consisted of five successive prolonged inflations. Sequential changes in clinical, intracoronary ECG and left ventricular indexes of myocardial ischemia were examined. Coronary collateral channels were evaluated during balloon inflations by ipsilateral and contralateral injections of contrast medium and hemodynamically by occlusion pressure. Results. An improved tolerance to myocardial ischemia with repetitive coronary occlusions was demonstrated by a significant reduction of angina, ST segment deviation, left ventricular filling pressure and less impairment of ejection fraction. Left ventricular wall motion abnormalities remained unchanged. Collateral angiographic grade did not change in 7 patients and increased in 10. Conclusions. This study confirms a progressive adaptation of myocardial ischemia to repetitive coronary occlusions and supports the concept that sequential episodes of myocardial ischemia are a stimulating factor for the recruitment of collateral channels in humans. These results also suggest that enhancement of recruitable collateral circulation might be an underlying mechanism of myocardial ischemic preconditioning.

A procedure is described for the disaggregation of colorectal cancers (CRC) and normal intestinal mucosal tissues to produce suspensions of viable single cells, which are then captured on customized antibody microarrays recognising 122 different surface antigens (DotScan CRC microarray). Cell binding patterns recorded by optical scanning of microarrays provide a surface profile of antigens on the cells. Sub-populations of cells bound on the microarray can be profiled by fluorescence multiplexing using monoclonal antibodies tagged with Quantum Dots or other fluorescent dyes. Surface profiles are presented for 6 CRC cell lines (T84, LIM1215, SW480, HT29, CaCo and SW620) and surgical samples from 40 CRC patients. Statistical analysis revealed significant differences between profiles for CRC samples and mucosal controls. Hierarchical clustering of CRC data identified several disease clusters that showed some correlation with clinico-pathological stage as determined by conventional histopathological analysis. Fluorescence multiplexing using Phycoerythrin- or Alexa Fluor 647-conjugated antibodies was more effective than multiplexing with antibodies labelled with Quantum Dots. This relatively simple method yields a large amount of information for each patient sample and, with further application, should provide disease signatures and enable the identification of patients with good or poor prognosis.

Purpose: As a pre-malignant precursor, adenoma provides an ideal tissue for proteome profiling to investigate early colorectal cancer development and provide possible targets for preventive interventions. The aim of this study was to identify patterns of differential protein expression that distinguish colorectal adenoma from normal tissue. Experimental design: Twenty paired samples of adenoma and normal mucosa were analysed by 2-DE and MALDI-TOF/TOF MS to detect proteins with ≥2-fold differential expression. Results: Four proteins were up-regulated in adenoma (Annexin A3, S100A11, S100P and eIF5A-1) and three were down-regulated (Galectin-1, S100A9 and FABPL). S100P, galectin-1, S100A9 and FABPL expression was localised by immunohistochemistry. Conclusions and clinical relevance: Distinctive patterns of in vivo protein expression in colorectal adenoma were identified for the first time. These proteins have important functions in cell differentiation, proliferation and metabolism, and may play a crucial role in early colorectal carcinogenesis. The ability to recognise premalignant lesions may have important applications in cancer prevention.

The significance of chromosome 3p gene alterations in lung cancer is poorly understood. This study set out to investigate promoter methylation in the deleted in lung and oesophageal cancer 1 (DLEC1), MLH1 and other 3p genes in 239 non-small cell lung carcinomas (NSCLC). DLEC1 was methylated in 38.7%, MLH1 in 35.7%, RARbeta in 51.7%, RASSF1A in 32.4% and BLU in 35.3% of tumours. Any two of the gene alterations were associated with each other except RARbeta. DLEC1 methylation was an independent marker of poor survival in the whole cohort (P=0.025) and in squamous cell carcinoma (P=0.041). MLH1 methylation was also prognostic, particularly in large cell cancer (P=0.006). Concordant methylation of DLEC1/MLH1 was the strongest independent indicator of poor prognosis in the whole cohort (P=0.009). However, microsatellite instability and loss of MLH1 expression was rare, suggesting that MLH1 promoter methylation does not usually lead to gene silencing in lung cancer. This is the first study describing the prognostic value of DLEC1 and MLH1 methylation in NSCLC. The concordant methylation is possibly a consequence of a long-range epigenetic effect in this region of chromosome 3p, which has recently been described in other cancers.

The aims of this study were to examine the associations between tumor adherence and other operative findings, postoperative complications, recurrence, and survival after resection of colorectal cancer.The prognostic importance of tumor adherence to other organs or structures, either by direct invasion (T4) or simply by inflammatory adhesions, is yet to be clearly defined as earlier studies have been limited in size or have not used contemporary multivariable statistical techniques.Data were drawn from a comprehensive, prospective hospital registry of all resections for colorectal cancer between January 1971 and December 2000 with follow-up to December 2005. Statistical analysis employed the chi test, Kaplan-Meier estimation, and proportional hazards regression with a significance level of0.05 and 95% confidence intervals (CI).Tumor adherence was identified in 268 of 2504 resections (10.7%). Adherent tumors were more likely than nonadherent tumors to be spontaneously or surgically perforated or transected, to have nodal metastases and to be poorly differentiated. Venous invasion was more frequent in adherent colonic but not rectal tumors. Adherence was associated with only 5 of 16 medical and surgical complications considered. In rectal cancer, adherence was independently associated with pelvic recurrence (hazard ratio 1.8, 95% CI 1.2-2.7) and diminished survival (hazard ratio 1.6, 95% CI 1.3-2.0) after adjustment for other variables.In rectal cancer, tumor adherence indicates a poor prognosis after adjustment for other prognostic factors, regardless of whether actual tumor invasion of the adherent structure has occurred. However, adherence is not associated with survival after resection of colonic cancer.