Immune related endonucleases and GTPases are not associated with tumor response in patients with advanced non-small cell lung cancer treated with checkpoint inhibitors.

Immune related endonucleases have recently been described as potential therapeutic targets and predictors of response to treatment with immune checkpoint inhibitors (ICI). The aim is to evaluate the association between the expression of 5 biomarkers involved in the immune response (CD73, CD39, VISTA, Arl4d and Cytohesin-3) in parallel with the more common ICI-predictive markers, PD-L1 expression and Tumor Mutation Burden (TMB) with response to ICI therapy in an advanced non-small cell lung cancer (NSCLC) cohort. METHODS: Patients with advanced NSCLC treated with ICI single agent were divided into responders and non-responders according to RECIST v1.1 and duration of response (DOR) criteria. Immunohistochemistry was performed on pretreatment tumor tissue samples for PD-L1, CD73, CD39, VISTA, Arl4d, and Cytohesin-3 expression. TMB was estimated with NEOplus v2 RUO (NEO New Oncology GmbH) hybrid capture next generation sequencing assay. Resistance mutations in STK11/KEAP1 and positive predictive mutations in ARID1A/POLE were also evaluated. RESULTS: Included were 56 patients who were treated with ICI single agent. The median progression-free and overall survival for the whole cohort was 3.0 (95% CI, 2.4-3.6) and 15 (95% CI, 9.7-20.2) months, respectively. The distribution of CD73 in tumor cells and CD39, VISTA, Arl4d and Cytohesin-3 expression in immune cells were not different between responders and non-responders. Also, PD-L1 and TMB were not predictive for response. The frequency of STK11, KEAP1 and ARID1A mutations was low and only observed in the non-responder group. CONCLUSION: Separate and combined expression of 5 biomarkers involved in the immune response (CD73, CD39, VISTA, Arl4d, and Cytohesin-3) was not associated with response in our cohort of advanced NSCLC patients receiving single agent ICI. To confirm our findings the analysis of independent larger cohorts is warranted.

Beneficial antioxidant status of piglets from sows fed selenomethionine compared with piglets from sows fed sodium selenite.

BACKGROUND: Studies in mammals proved dietary organic selenium (Se) being superior to inorganic Se regarding effects on growth performance, antioxidative status, immune response, and Se homeostasis. However, the picture of possible effects of different Se sources and - levels can be expanded. The present field study evaluated the effects on weight gain, hematological and selected biochemical variables as well as plasma concentrations of vitamin E (vitE), total Se and selenobiomolecules in piglets throughout the suckling period. METHODS: Piglets were monitored from birth to 38 days of age (d). The mother sows' diets were enriched with l-selenomethionine (SeMet-0.26 and -0.43 mg Se/kg feed) or sodium selenite (NaSe-0.40 and -0.60 mg Se/kg feed) from 1 month prior to farrowing until the end of lactation period. Piglets received pelleted feed supplemented with Se similarly to the sows' diets from one week of age. Selenite at 0.40 mg Se/kg (NaSe-0.40) represents a common Se source and -level in pig feed and served as control diet. RESULTS: From 24d, piglets in SeMet-groups had higher mean body weight (BW) compared with piglets from sows fed NaSe-0.40. Furthermore, from five-d and above, piglets from sows fed NaSe-0.60 had significantly higher BW than offspring from sows fed NaSe-0.40. Neonatal piglets in group SeMet-0.43 had significantly lower red blood cell counts (RBC), hemoglobin (Hgb) and hematocrit (Hct) concentrations compared with piglets from sows fed with NaSe-0.40. Neonatal and 5d-old piglets in group SeMet-0.26 showed higher gamma-glutamyl transferase activity than piglets in group NaSe-0.40. From five d and above, group NaSe-0.60 excelled with increased specific hematological variables culminating at age 38d with increased Hct, mean corpuscular volume (MCV), and MC hemoglobin (MCH) as well as increased activities of aspartate transaminase and lactate dehydrogenase compared with the other groups. Generally, offspring in the SeMet groups had higher total Se-concentrations in plasma than those from sows fed selenite, and showed a dose-response effect on plasma Se-concentrations. Furthermore, SeMet-fed piglets had higher plasma levels of the selenoproteins (Sel) glutathione peroxidase 3 (GPx3) and SelP as well as selenoalbumin. Plasma vitE levels were significantly negatively correlated with RBC throughout trial period. CONCLUSIONS: Maternal supplementation with SeMet during gestation influenced hematology and clinical biochemistry in neonatal piglets in a different way than in offspring from sows receiving selenite enriched diets. Growth performance was positively influenced by both dietary Se source and Se level. Higher plasma levels of GPx3 observed in piglets receiving SeMet probably improved the protection against birth or growth related oxidative stress. These might prime the piglets for demanding situations as indicated by higher weight gain in offspring from sows fed with SeMet-supplemented diets. Our results on some enzyme activities might indicate that piglets fed NaSe-0.60 had to cope with increased levels of oxidative stress compared with those originating from sows fed SeMet or lower dietary levels of selenite. We assume that combining inorganic and organic Se sources in complete feed for breeding sows might be beneficial fro reproduction and the offspring's performance.

An independent assessment of reliability of the Melbourne Cerebral Palsy Hip Classification System.

PURPOSE: Neuromuscular (NM) hip dysplasia is common in patients with cerebral palsy (CP). Traditionally, migration percentage (MP) has been used to measure the severity of NM hip dysplasia; however, the MP has some limitations. The purpose of this study is to determine the intra- and inter-reliability of the Melbourne Cerebral Palsy Hip Classification System in the typical paediatric population of patients with CP. METHODS: A total of 65 anteroposterior pelvis radiographs in patients (age range 12 years to 21 years) with CP spanning all grades (I to VI) of the classification system were identified and collected for analysis in this institutional review board approved study. Four paediatric orthopaedic surgeons and one orthopaedic surgical resident classified each radiograph according to the Melbourne system. Then, at least four weeks later, the raters repeated the process with a re-randomised order of radiographs. Statistical analysis was performed using the intraclass correlation coefficient (ICC) where < 0 denotes poor agreement and > 0.8 indicates almost perfect agreement. RESULTS: The interobserver reliability was found to be excellent with the ICC of 0.853 (0.813 to 0.887) and 0.839 (0.795 to 0.877). The intraobserver reliability was also found to be excellent with the ICC in the range of 0.838 to 0.933 among the raters. Subgroup analysis indicated no differences in the reliability of observers based on clinical experience. CONCLUSION: This study independently demonstrates that the Melbourne Cerebral Palsy Hip Classification System for NM hip dysplasia in patients with CP can be reliably used for communication among various healthcare providers and research and epidemiological purposes.

Prognostic risk factors of first recurrence in patients with primary stages I-II cutaneous malignant melanoma - from the population-based Swedish melanoma register.

BACKGROUND: Prognostic factors in patients with localized primary cutaneous malignant melanoma (CMM) are well described. However, prognostic factors for recurrence are less documented. OBJECTIVES: The aim of this study was to identify prognostic risk factors for first recurrence in patients with localized stages I-II CMM using population-based data. METHODS: This study included 1437 CMM patients registered in one region of Sweden during 1999-2012 follow-up through 31 December 2012. To identify first recurrence of CMM disease, data from a care data warehouse, the pathology and radiology department registries were used. Patients were also followed through a census register and the national Cause of Death Register. RESULTS: The 5- and 10-year recurrence-free survival (RFS) were 85.7% and 81.2%, respectively. The most common site of first recurrence was regional lymph node metastasis closely followed by distant metastasis. After adjusting for all prognostic factors, women had 50% lower risk of recurrence than men (HR = 0.5, 95% CI 0.4-0.7) and patients ≥70 had higher risk compared to patients 55-69 years (HR = 1.7, 95% CI 1.2-2.5). Other significant prognostic factors for risk of recurrence were tumour thickness, presence of ulceration, Clark's level of invasion and histogenetic type. CONCLUSION: Tumour thickness was found to be the predominant risk factor for recurrence. The prognostic factors for recurrence coincided with prognostic factors for CMM death. The most common site of first recurrence in stages I-II CMM is regional lymph node (42.8%) closely followed by distant metastases (37.6%), a fact which has to be taken into consideration when choosing follow-up strategies.

Fundoplication for laryngopharyngeal reflux despite preoperative dysphagia.

INTRODUCTION Fundoplication for laryngopharyngeal disease with oesophageal dysmotility has led to mixed outcomes. In the presence of preoperative dysphagia and oesophageal dysmotility, this procedure has engendered concern in certain regards. METHODS This paper describes a consecutive series of laryngopharyngeal reflux (LPR) patients with a high frequency of dysmotility. Patients were selected for surgery with 24-hour dual channel pH monitoring, oesophageal manometry and standardised reflux scintigraphy. RESULTS Following careful patient selection, 33 patients underwent fundoplication by laparoscopy. Surgery had high efficacy in symptom control and there was no adverse dysphagia. CONCLUSIONS Evidence of proximal reflux can select a group of patients for good results of fundoplication for atypical symptoms.

Validation of sun exposure and protection index (SEPI) for estimation of sun habits.

BACKGROUND: In both Sweden and Australia high incidence rates of skin cancer have become a major health problem. In prevention and risk communication, it is important to have reliable ways for identifying people with risky sun habits. In this study the validity and reliability of the questionnaire Sun Exposure Protection Index (SEPI), developed to assess individual's sun habits and their propensity to increase sun protection during routine, often brief, clinical encounters, has been evaluated. The aim of our study was to evaluate validity and reliability of the proposed SEPI scoring instrument, in two countries with markedly different ultraviolet radiation environments (Sweden and Australia). METHOD: Two subpopulations in Sweden and Australia respectively were asked to fill out the SEPI together with the previously evaluated Readiness to Alter Sun Protective Behaviour questionnaire (RASP-B) and the associated Sun-protective Behaviours Questionnaire. To test reliability, the SEPI was again filled out by the subjects one month later. RESULTS: Comparison between SEPI and the questions in the Sun-protective Behaviours Questionnaire, analyzed with Spearman's Rho, showed good correlations regarding sun habits. Comparison between SEPI and RASP-B regarding propensity to increase sun protection showed concurrently lower SEPI mean scores for action stage, but no difference between precontemplation and contemplation stages. The SEPI test-retest analysis indicated stability over time. Internal consistency of the SEPI, assessed with Cronbach's alpha estimation showed values marginally lower than the desired >0.70 coefficient value generally recommended, and was somewhat negatively affected by the question on sunscreen use, likely related to the classic "sunscreen paradox". There were some differences in the performance of the SEPI between the Swedish and Australian samples, possibly due to the influence of "available" sunlight and differing attitudes to behaviour and protection "at home" and on vacation. CONCLUSIONS: SEPI appears to be a stable instrument with an overall acceptable validity and reliability, applicable for use in populations exposed to different UVR environments, in order to evaluate individual sun exposure and protection.

Near infrared and skin impedance spectroscopy - a possible support in the diagnostic process of skin tumours in primary health care.

BACKGROUND/PURPOSE: The global incidence of skin cancer has increased drastically in recent decades, especially in Australia and Northern Europe. Early detection is crucial for good prognosis and high survival rates. In general, primary care physicians have considerably lower sensitivity and specificity rates for detection of skin cancer, compared to dermatologists. A probable main reason for this is that current diagnostic tools are subjective in nature, and therefore diagnostic skills highly depend on experience. Illustratively, in Sweden, approximately 155 500 benign skin lesions are excised unnecessarily every year. An objective instrument, added to the clinical examination, might improve the diagnostic accuracy, and thus promote earlier detection of malignant skin tumours, as well as reduce medical costs associated with unnecessary biopsies and excisions. The general aim of this study was to investigate the usefulness of the combination of near infrared (NIR) and skin impedance spectroscopy as a supportive tool in the diagnosis and evaluation of skin tumours in primary health care. METHODS: Near infrared and skin impedance data were collected by performing measurements on suspect malignant, premalignant and benign tumours in the skin of patients seeking primary health care for skin tumour evaluation. The obtained data were analysed using multivariate analysis and compared with the diagnosis received by the conventional diagnostic process. RESULTS: The observed sensitivity and specificity rates were both 100%, when discriminating malignant and premalignant skin tumours from benign skin tumours, and the observed sensitivity and specificity for separating malignant skin tumours from premalignant and benign skin tumours were also 100%, respectively. CONCLUSION: The results of this study indicate that the NIR and skin impedance spectroscopy may be a useful supportive tool for the general practitioner in the diagnosis and evaluation of skin tumours in primary health care, as a complement to the visual assessment.

Tecemotide in unresectable stage III non-small-cell lung cancer in the phase III START study: updated overall survival and biomarker analyses.

BACKGROUND: Tecemotide is a MUC1-antigen-specific cancer immunotherapy. The phase III START study did not meet its primary end point but reported notable survival benefit with tecemotide versus placebo in an exploratory analysis of the predefined patient subgroup treated with concurrent chemoradiotherapy. Here, we attempted to gain further insight into the effects of tecemotide in START. PATIENTS AND METHODS: START recruited patients who did not progress following frontline chemoradiotherapy for unresectable stage III non-small-cell lung cancer. We present updated overall survival (OS) data and exploratory analyses of OS for baseline biomarkers: soluble MUC1 (sMUC1), antinuclear antibodies (ANA), neutrophil/lymphocyte ratio (NLR), lymphocyte count, and HLA type. RESULTS: Updated OS data are consistent with the primary analysis: median 25.8 months (tecemotide) versus 22.4 months (placebo) (HR 0.89, 95% CI 0.77-1.03, P = 0.111), with ∼20 months additional median follow-up time compared with the primary analysis. Exploratory analysis of the predefined subgroup treated with concurrent chemoradiotherapy revealed clinically relevant prolonged OS with tecemotide versus placebo (29.4 versus 20.8 months; HR 0.81, 95% CI 0.68-0.98, P = 0.026). No improvement was seen with sequential chemoradiotherapy. High sMUC1 and ANA correlated with a possible survival benefit with tecemotide (interaction P = 0.0085 and 0.0022) and might have future value as biomarkers. Interactions between lymphocyte count, NLR, or prespecified HLA alleles and treatment effect were not observed. CONCLUSION: Updated OS data support potential treatment benefit with tecemotide in patients treated with concurrent chemoradiotherapy. Exploratory biomarker analyses suggest that elevated sMUC1 or ANA levels correlate with tecemotide benefit. CLINICALTRIALSGOV NUMBER: NCT00409188.

Frequent chromatin rearrangements in myelodysplastic syndromes--what stands behind?

Myelodysplastic syndromes (MDS) represent a clinically and genetically heterogeneous group of clonal haematopoietic diseases characterized by a short survival and high rate of transformation to acute myeloid leukaemia (AML). In spite of this variability, MDS is associated with typical recurrent non-random cytogenetic defects. Chromosomal abnormalities are detected in the malignant bone-marrow cells of approximately 40-80 % of patients with primary or secondary MDS. The most frequent chromosomal rearrangements involve chromosomes 5, 7 and 8. MDS often shows presence of unbalanced chromosomal changes, especially large deletions [del(5), del(7q), del(12p), del(18q), del(20q)] or losses of whole chromosomes (7 and Y). The most typical cytogenetic abnormality is a partial or complete deletion of 5q- that occurs in roughly 30 % of all MDS cases either as the sole abnormality or in combination with other aberrations as a part of frequently complex karyotypes. The mechanisms responsible for the formation of MDS-associated recurrent translocations and complex karyotypes are unknown. Since some of the mentioned aberrations are characteristic for several haematological malignancies, more general cellular conditions could be expected to play a role. In this article, we introduce the most common rearrangements linked to MDS and discuss the potential role of the non-random higher-order chromatin structure in their formation. A contribution of the chromothripsis - a catastrophic event discovered only recently - is considered to explain how complex karyotypes may occur (during a single event).

Influence of age, gender, educational level and self-estimation of skin type on sun exposure habits and readiness to increase sun protection.

BACKGROUND: Sun exposure habits and the propensity to undertake sun protection differ between individuals. Not least in primary prevention of skin cancer, aiming at reducing ultraviolet (UV) exposure, knowledge about these factors may be of importance. The aim of the present study was to investigate, in a primary health care (PHC) population, the relationship between sun exposure habits/sun protection behaviour/readiness to increase sun protection and gender, age, educational level and skin UV-sensitivity. METHODS: The baseline data from a previously performed RCT on skin cancer prevention was used. 415 patients, aged > 18 years, visiting a PHC centre in southern Sweden, filled-out a questionnaire mapping sun exposure, readiness to increase sun protection according to the Transtheoretical Model of Behaviour Change (TTM), and the above mentioned factors. RESULTS: Female gender was associated with more frequent suntanning (p < 0.001) and sunbed use (p < 0.05), but also with more extensive sunscreen use (p < 0.001). High age was in general associated with low level of sun exposure and high level of protection. Subjects with low educational level reported less frequent sunscreen use than those with higher educational level, and also chose lower SPF (p < 0.001). For almost all parameters, high skin UV-sensitivity was associated with markedly lower sun exposure (p < 0.001) and more pronounced readiness to increase sun protection. Females and subjects with high educational level reported higher readiness to increase sunscreen use than males and subjects with lower educational level (p < 0.001). CONCLUSIONS: Gender, age, educational level and skin type appear to be important factors affecting sun exposure habits and sun protection behaviour, which supports the idea of appropriate mapping of these factors in patients in order to individualise sun protection advice according to the individual patient situation and capabilities.

Measuring sun exposure habits and sun protection behaviour using a comprehensive scoring instrument--an illustration of a possible model based on Likert scale scorings and on estimation of readiness to increase sun protection.

BACKGROUND: Few attempts to present a comprehensive scoring instrument for sun exposure and protection have been made. The present paper aims to describe a possible set of questions suitable for such an instrument, comprising the most important aspects of sun exposure and protection. METHODS: The material from a previously performed intervention study, using a questionnaire based on Likert scales and on the Transtheoretical Model of Behaviour Change (TTM), was utilised. 213 primary healthcare patients filled in the questionnaire and were randomised into two groups receiving sun protection advice, in Group 1 in letter-form, and in Group 2 orally during a doctor's consultation. In the original study, increased sun protection/readiness to increase sun protection was demonstrated for several items in Group 2, at six months. To compose a comprehensive scoring instrument, five questions concerning sun exposure/protection (intentional tanning, sunscreen use, choice of SPF, number of occasions with sunburn, and time spent in the sun at midday), were selected to give a 20 point behavioural score. Similarly, four TTM-based questions (giving up sunbathing, using clothes for sun protection, using sunscreens, and staying in the shade) gave a 16 point "propensity-to-change"-score. RESULTS: At follow-up, increased sun protection reflected in the behavioural score occurred only in Group 2 (p < 0.001). For the propensity-to-change-score, increased readiness to increase sun protection occurred in both groups, but the change was significantly higher in Group 2 (p < 0.05). Categorisation of the 20 point behavioural score, into three risk levels, revealed a significantly higher shift of subjects moving to a lower risk level in Group 2 compared to Group 1 (p<0.05). CONCLUSIONS: In conclusion, twinning of a summarised Likert scale behavioural score with a TTM-based propensity-to-change-score seems promising for the creation of a questionnaire-based, comprehensive scoring instrument for sun exposure and protection.

Evaluation of 3D nano-macro porous bioactive glass scaffold for hard tissue engineering.

Recently, nano-macro dual-porous, three-dimensional (3D) glass structures were developed for use as bioscaffolds for hard tissue regeneration, but there have been concerns regarding the interconnectivity and homogeneity of nanopores in the scaffolds, as well as the cytotoxicity of the environment deep inside due to limited fluid access. Therefore, mercury porosimetry, nitrogen absorption, and TEM have been used to characterize nanopore network of the scaffolds. In parallel, viability of MG 63 human osteosarcoma cells seeded on scaffold surface was investigated by fluorescence, confocal and electron microscopy methods. The results show that cells attach, migrate and penetrate inside the glass scaffold with high proliferation and viability rate. Additionally, scaffolds were implanted under the skin of a male New Zealand rabbit for in vivo animal test. Initial observations show the formation of new tissue with blood vessels and collagen fibers deep inside the implanted scaffolds with no obvious inflammatory reaction. Thus, the new nano-macro dual-porous glass structure could be a promising bioscaffold for use in regenerative medicine and tissue engineering for bone regeneration.

Etanercept treatment of renal amyloidosis complicating rheumatoid arthritis.

Rheumatoid arthritis and juvenile arthritis represent the commonest diseases complicated by AA amyloidosis in developed countries. Up to 5% of patients with rheumatoid arthritis will develop AA amyloidosis, with renal failure being the commonest cause of mortality. To date, treatment of this condition has focused on suppressing the underlying inflammatory condition with drugs such as cyclophosphamide and chlorambucil, but both these drugs are associated with myelotoxicity, leukaemia and sterility. Tumour necrosis factor-alpha (TNF-alpha) is thought to be involved in amyloid deposition. The efficacy of anti-TNF-alpha therapy (etanercept) in the treatment of renal amyloidosis complicating rheumatoid arthritis is demonstrated here and the current scientific data on this subject are presented.

Neoadjuvant chemotherapy combined with regional hyperthermia (RHT) for locally advanced primary or recurrent high-risk adult soft-tissue sarcomas (STS) of adults: long-term results of a phase II study.

In this phase II study, activity and safety of neoadjuvant regional hyperthermia (RHT) combined with chemotherapy was investigated in 59 patients with primary advanced or recurrent high-risk soft-tissue sarcoma (STS). Patients received four EIA cycles consisting of etoposide, ifosfamide and doxorubicin combined with RHT followed by surgical resection and adjuvant treatment. The overall objective response (OR) rate was 17%, with one complete (2%) and eight partial (15%) responses. In addition, 13 minor responses (25%) were seen. At time of surgery, complete necrosis (pCR) occurred in 6 patients and >75% necrosis (favourable histological response (FHR)) in 12 patients. At the completion of protocol treatment, 36 patients were rendered disease-free which was significantly associated with the initial radiographic and/or pathological tumour response (P=0.004). Treatment-related toxicity was acceptable overall. At a medium follow-up of 82 months, local treatment failure occurred in 33 patients, median overall survival (OS) was 52 months, and the 5-year survival rate was 49% (95% confidence interval (CI): 36-61%). OS which did not differ for extremity versus non-extremity STS (P=0.21) was better for patients responding to EIA combined with RHT (P<0.01).

Treatment of primary, recurrent or inadequately resected high-risk soft-tissue sarcomas (STS) of adults: results of a phase II pilot study (RHT-95) of neoadjuvant chemotherapy combined with regional hyperthermia.

The efficacy of thermochemotherapy in adult patients with primary, recurrent or inadequately resected non-metastatic high-risk soft-tissue sarcomas (STS) was assessed. 54 patients were prospectively treated with four cycles of etoposide, ifosfamide and doxorubicin (EIA) combined with regional hyperthermia (RHT) followed by surgery, another four cycles of EIA without RHT and external beam radiation. The objective response rate was 16% and at a median follow-up time of 57 months, the 4-year estimated rates of local failure-free survival (LFFS), distant metastasis-free survival (DMFS), event-free survival (EFS) and overall survival (OS) were 59% (95% confidence interval (CI) 45-73%), 59% (95% CI 44-73%), 26% (95% CI 14-38%) and 40% (95% CI 27-53%), respectively. OS was in favour of patients responding to neoadjuvant treatment (P=0.073). In comparison to a preceding phase II study including pre- and postsurgical thermochemotherapy (RHT-91), at a 4-year follow-up the RHT-95 study cohort showed an inferior LFFS rate (P=0.027), but this did not affect DMFS (P=0.558) or OS (P=0.126). Hence, postsurgical thermochemotherapy seems critical for local tumour control without affecting survival.

High resolution, fluorescence deconvolution microscopy and tagging with the autofluorescent tracers CFP, GFP, and YFP to study the structural composition of gap junctions in living cells.

High-resolution, fluorescence deconvolution (DV) microscopy was implemented to obtain a detailed view of the organization and structural composition of gap junctions assembled from one or two different connexin isotypes in live and fixed cells. To visualize gap junctions, the structural protein components of gap junction channels, the connexin polypeptides alpha1(Cx43), beta1(Cx32), and beta2(Cx26), were tagged on their C-termini with the autofluorescent tracers green fluorescent protein (GFP), and its cyan (CFP), and yellow (YFP) color variants. Tagged connexins were expressed in transiently transfected HeLa cells. Comprehensive analysis including dye-transfer analysis demonstrated that the tagged connexins trafficked, assembled, and packed normally into functional gap junction channel plaques. Such gap junction plaques were examined by single, dual, and triple-color DV microscopy. High-resolution images and three-dimensional volume reconstructions of gap junction plaques were obtained by this technique, which revealed several new aspects of gap junction structure. Specifically, the studies demonstrated that the mode of channel distribution strictly depends on the connexin isotypes. Here we present such images, and volume reconstructions in context with images obtained by other light, and electron microscopic techniques, such as laser scanning confocal, conventional wide-field fluorescence, thin section, and freeze-fracture electron microscopy. In addition, we give a simple description of the principal mechanisms of DV microscopy, name advantages and disadvantages, and discuss issues such as dual-color imaging using CFP and YFP, spatial resolution, colocalization, and avoiding imaging artifacts.

Hyperthermia in oncology.

The purpose of this article is to provide an overview on the current clinical application of hyperthermia combined with conventional treatment modalities (e.g. ionizing radiation, chemotherapy) in the treatment of malignant disease. The clinical application of hyperthermia with increase of tissue temperatures (range 40-44 degrees C) has been integrated in multimodal anti-cancer strategies. This review describes selected phase I or II (n = 17) and phase III trials (n = 16) investigating the effect of hyperthermia combined with radiotherapy (n = 10 trials), chemotherapy (n = 15 trials), or both (n = 8 trials) in a total of more than 2200 patients. The trials were performed in a variety of solid tumours (e.g. melanoma, head and neck cancer, breast cancer, cancer of the gastrointestinal or urogenital tract, glioblastoma, sarcoma) in paediatric or adult patients. Profound research has produced a scientific basis for the simultaneous application of hyperthermia in combination with ionizing radiation and/or systemic chemotherapy. Hyperthermia is becoming more accepted clinically, due to the substantial technical improvements made in achieving selected increase of temperatures in superficial and deep-seated tumours. At present, the combination of hyperthermia and chemotherapy or radiochemotherapy is further tested within clinical protocols (phase II/III) in order to improve local tumour control and relapse-free survival in patients with high-risk or advanced tumours of different entities.

Biosynthesis and structural composition of gap junction intercellular membrane channels.

Gap junction channels assemble as dodecameric complexes, in which a hexameric connexon (hemichannel) in one plasma membrane docks end-to-end with a connexon in the membrane of a closely apposed cell to provide direct cell-to-cell communication. Synthesis, assembly, and trafficking of the gap junction channel subunit proteins referred to as connexins, largely appear to follow the general secretory pathway for membrane proteins. The connexin subunits can assemble into homo-, as well as distinct hetero-oligomeric connexons. Assembly appears to be based on specific signals located within the connexin polypeptides. Plaque formation by the clustering of gap junction channels in the plane of the membrane, as well as channel degradation are poorly understood processes that are topics of current research. Recently, we tagged connexins with the autofluorescent reporter green fluorescent protein (GFP), and its cyan (CFP), and yellow (YFP) color variants and combined this reporter technology with single, and dual-color, high resolution deconvolution microscopy, computational volume rendering, and time-lapse microscopy to examine the detailed organization, structural composition, and dynamics of gap junctions in live cells. This technology provided for the first time a realistic, three-dimensional impression of gap junctions as they appear in the plasma membranes of adjoining cells, and revealed an excitingly detailed structural organization of gap junctions never seen before in live cells. Here, I summarize recent progress in areas encompassing the synthesis, assembly and structural composition of gap junctions with a special emphasis on the recent results we obtained using cell-free translation/ membrane-protein translocation, and autofluorescent reporters in combination with live-cell deconvolution microscopy.

Novel surveillance and cure of a donor-transmitted lymphoma in a renal allograft recipient.

BACKGROUND: In this report we describe a malignant lymphoma of donor origin inadvertently transplanted into two renal allograft recipients, despite standard comprehensive donor screening. The successful clearance of the tumor from both patients and a novel method of surveillance are detailed. METHODS: Initial management consisted of withdrawal of immunosuppression to promote rejection of the allograft and the transplanted tumor in both patients, followed by graft removal. Peripheral blood microchimerism was assessed in both recipients using nested polymerase chain reaction to detect the DYZ3 gene on the Y chromosome (donor male, recipients female). RESULTS: Although microchimerism was detected on day 6 after transplantation and day 1 after explantation, repeat peripheral blood examination at 1, 3, and 6 months after explantation demonstrated no microchimerism. Both patients remain well at 12 months and have been relisted for transplantation. CONCLUSION: Despite inadvertent transplantation of a previously undiagnosed malignancy of donor origin, the recipients' immune response was able to eliminate donor tumor cells after the withdrawal of immunosuppression. Repeated surveillance of peripheral blood from both recipients, using a novel application of the technique of nested polymerase chain reaction to amplify donor DNA, demonstrated no persistence of donor cells, supporting effective eradication of the donor malignancy.