Circulating miRNAs in Women with Polycystic Ovary Syndrome: A Longitudinal Cohort Study.

BACKGROUND: Women with polycystic ovary syndrome (PCOS) often change their metabolic profile over time to decrease levels of androgens while often gaining a propensity for the development of the metabolic syndrome. Recent discoveries indicate that microRNAs (miRNAs) play a role in the development of PCOS and constitute potential biomarkers for PCOS. We aimed to identify miRNAs associated with the development of an impaired metabolic profile in women with PCOS, in a follow-up study, compared with women without PCOS. METHODS AND MATERIALS: Clinical measurements of PCOS status and metabolic disease were obtained twice 6 years apart in a cohort of 46 women with PCOS and nine controls. All participants were evaluated for degree of metabolic disease (hypertension, dyslipidemia, central obesity, and impaired glucose tolerance). MiRNA levels were measured using Taqman® Array cards of 96 pre-selected miRNAs associated with PCOS and/or metabolic disease. RESULTS: Women with PCOS decreased their levels of androgens during follow-up. Twenty-six of the miRNAs were significantly changed in circulation in women with PCOS during the follow-up, and twenty-four of them had decreased, while levels did not change in the control group. Four miRNAs were significantly different at baseline between healthy controls and women with PCOS; miR-103-3p, miR-139-5p, miR-28-3p, and miR-376a-3p, which were decreased in PCOS. After follow-up, miR-28-3p, miR-139-5p, and miR-376a-3p increased in PCOS women to the levels observed in healthy controls. Of these, miR-139-5p correlated with total testosterone levels (rho = 0.50, padj = 0.013), while miR-376-3p correlated significantly with the waist-hip ratio at follow-up (rho = 0.43, padj = 0.01). Predicted targets of miR-103-3p, miR-139-5p, miR-28-3p, and miR-376a-3p were enriched in pathways associated with Insulin/IGF signaling, interleukin signaling, the GNRH receptor pathways, and other signaling pathways. MiRNAs altered during follow-up in PCOS patients were enriched in pathways related to immune regulation, gonadotropin-releasing hormone signaling, tyrosine kinase signaling, and WNT signaling. CONCLUSIONS: These studies indicate that miRNAs associated with PCOS and androgen metabolism overall decrease during a 6-year follow-up, reflecting the phenotypic change in PCOS individuals towards a less hyperandrogenic profile.

Efficacy of mesenchymal stem cell-delivery using perpendicular multi-needle injections to the skin: Evaluation of post-ejection cellular health and dermal delivery.

AIM: Mesenchymal stem cell (MSC)-therapy is increasingly being evaluated in clinical trials. Dermal delivery is not only time consuming but also unreliable, potentially hampering the therapeutic result. Therefore, qualification of cell delivery protocols is essential. This study evaluated a clinically relevant automated multi-needle injection method for cutaneous MSC-therapy, allowing the skin to be readily and timely treated, by assessing both the cellular health post-ejection and dermal delivery. METHODS: Following dispensation through the injector (31 G needles: 9- or 5-pin) the cellular health and potency (perceived- and long-term (12 h) viability, recovery, metabolism, adherence, proliferation and IDO1-expression) of adipose-derived stem cells (10-20-50 ×106 cells/ml) were assessed in vitro in addition to dermal delivery of solution in human skin. RESULTS: No significant detrimental effect on the perceived cell viability, recovery, metabolism, adherence or IDO1-expression of either cell concentration was observed. However, the overall long-term viability and proliferation decreased significantly regardless of cell concentration, nonetheless marginally. An injection depth above 1.0 mm resulted in all needles piercing the skin with dermal delivery from up to 89% needles and minimal reflux to the skin surface, and the results were confirmed by ultrasound and histology. CONCLUSION: The automated injector is capable of delivering dermal cell-doses with an acceptable cell quality.

Effect of weight and storage time of broiler breeders' eggs on morphology and biochemical features of eggs, embryogenesis, hatchability, and chick quality.

The transfer of hatchability results obtained under experimental conditions to the commercial ground with a positive financial effect proves the value and usefulness of these data. On the other hand, finding results on commercial processes of broiler breeders' egg incubation in the literature is challenging. The presented study aimed to determine the effects of egg weight and storage time on the physical, biochemical characteristics of hatching eggs, embryogenesis and hatchability in Ross 308 broiler breeders. On the laying day, the eggs were divided into four weight groups: S - small eggs (57-61 g), M - medium eggs (62-66 g), L - large eggs (67-71 g), and XL - extra-large eggs (72-76 g). The eggs were then stored for 3, 7, 14, and 21 days under controlled conditions. As the egg storage time increased, a decrease in the yolk quality (lower index) was observed. The highest Haugh units were found in eggs from the S and M groups. The cholesterol content of the M, L, and XL groups was lower on days 7, 14, and 21 as compared to that of eggs only stored for 3 days. Egg weight loss during incubation decreased with an increase in the egg weight. An extension of the egg storage time caused an increase in the loss of egg weight. On the 14th and 18th days of hatching, an increase in the eggshell temperature was noted with an increase in the weight of the egg. The eggs stored for 7 days were characterised by the highest shell temperature on each day. The highest hatchability percentage was recorded for the M group. The hatchability rate decreased with the prolongation of the storage time, while the number of crippled chicks after hatching increased. The results confirmed that the increased weight of the eggs and prolonged storage time (14 and 21 days) increased the weight and decreased the length of the newly hatched chicks, respectively. Chicks from the heaviest eggs and those stored for 14 and 21 days showed poor results on the Pasgar score® test. The observations indicate the need to adopt various (of those available) methods to assess the quality of newly hatched chicks in hatcheries in order to produce high-quality broiler chickens. The results also indicate that prolonged egg storing beyond 14 days may affect the thyroid hormone economy during the hatching of chicks, especially in the XL group.

HLA Class II Histocompatibility Antigen γ Chain (CD74) Expression Is Associated with Immune Cell Infiltration and Favorable Outcome in Breast Cancer.

The triple-negative breast cancer (TNBC) subtype, defined as negative for ER, PgR, and HER2, is biologically more aggressive and with a poorer prognosis than the other subtypes, in part due to the lack of suitable targeted therapies. Consequently, identification of any potential novel therapeutic option, predictive and/or prognostic biomarker, or any other relevant information that may impact the clinical management of this group of patients is valuable. The HLA class II histocompatibility antigen γ chain, or cluster of differentiation 74 (CD74), has been associated with TNBCs, and poorer survival. However, discordant results have been reported for immunohistochemical studies of CD74 expression in breast cancer. Here we report validation studies for use of a novel CD74 antibody, UMAb231. We used this antibody to stain a TMA including 640 human breast cancer samples, and found no association with the TNBC subtype, but did find a positive correlation with outcome. We also found associations between CD74 expression and immune cell infiltration, and expression of programmed death ligand 1 (PD-L1). Given that CD74 may play a role in innate immune system responses and the potential of immunotherapy as a viable treatment strategy for TNBCs, CD74 expression may have predictive value for immune checkpoint therapies.

Characterization and Outcomes of SARS-CoV-2 Infection in Patients with Sarcoidosis.

To analyze the clinical characteristics and outcomes of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in patients with sarcoidosis from a large multicenter cohort from Southern Europe and to identify the risk factors associated with a more complicated infection. We searched for patients with sarcoidosis presenting with SARS-CoV-2 infection (defined according to the European Centre for Disease Prevention and Control guidelines) among those included in the SarcoGEAS Registry, a nationwide, multicenter registry of patients fulfilling the American Thoracic Society/European Respiratory Society/World Association of Sarcoidosis and Other Granulomatous Disorders 1999 classification criteria for sarcoidosis. A 2:1 age-sex-matched subset of patients with sarcoidosis without SARS-CoV-2 infection was selected as control population. Forty-five patients with SARS-CoV-2 infection were identified (28 women, mean age 55 years). Thirty-six patients presented a symptomatic SARS-CoV-2 infection and 14 were hospitalized (12 required supplemental oxygen, 2 intensive care unit admission and 1 mechanical ventilation). Four patients died due to progressive respiratory failure. Patients who required hospital admission had an older mean age (64.9 vs. 51.0 years, p = 0.006), a higher frequency of baseline comorbidities including cardiovascular disease (64% vs. 23%, p = 0.016), diabetes mellitus (43% vs. 13%, p = 0.049) and chronic liver/kidney diseases (36% vs. 0%, p = 0.002) and presented more frequently fever (79% vs. 35%, p = 0.011) and dyspnea (50% vs. 3%, p = 0.001) in comparison with patients managed at home. Age- and sex-adjusted multivariate analysis identified the age at diagnosis of SARS-Cov-2 infection as the only independent variable associated with hospitalization (adjusted odds ratio 1.18, 95% conficence interval 1.04-1.35). A baseline moderate/severe pulmonary impairment in function tests was associated with a higher rate of hospitalization but the difference was not statistically significant (50% vs. 23%, p = 0.219). A close monitoring of SARS-CoV-2 infection in elderly patients with sarcoidosis, especially in those with baseline cardiopulmonary diseases and chronic liver or renal failure, is recommended. The low frequency of severe pulmonary involvement in patients with sarcoidosis from Southern Europe may explain the weak prognostic role of baseline lung impairment in our study, in contrast to studies from other geographical areas.

Different T cell related immunological profiles in COVID-19 patients compared to healthy controls.

In various pathological conditions, cellular immunity plays an important role in immune responses. Amongimmunecells, T lymphocytes pdomotecellular and humoralresponses as well as innate immunity. Therefore, careful investigation of these cells has a significant impact on accurate knowledge in COVID-19diseasepathogenesis. In current research, the frequency and function of various T lymphocytes involved in immune responses examined in SARS-CoV-2 patients with various disease severity compared to normal subjects. In order to make an accurate comparison among patients with various disease severity, this study was performed on asymptomatic recovered cases (n = 20), ICU hospitalized patients (n = 30), non-ICU hospitalized patients (n = 30), and normal subjects (n = 20). To precisely evaluate T cells activity following purification, their cytokine secretion activity was examined. Similarly, immediately after purification of Treg cells, their inhibitory activity on T cells was investigated. The results showed that COVID-19 patients with severe disease (ICU hospitalized patients) not only had a remarkable increase in Th1 and Th17 but also a considerable decrease in Th2 and Treg cells. More importantly, as the IL-17 and IFN-γ secretion was sharply increased in severe disease, the secretion of IL-10 and IL-4 was decreased. Furthermore, the inhibitory activity of Treg cells was reduced in severe disease patients in comparison to other groups. In severe COVID-19 disease, current findings indicate when the inflammatory arm of cellular immunity is significantly increased, a considerable reduction in anti-inflammatory and regulatory arm occurred.

A translational program that suppresses metabolism to shield the genome.

Translatome reprogramming is a primary determinant of protein levels during stimuli adaptation. This raises the question: what are the translatome remodelers that reprogram protein output to activate biochemical adaptations. Here, we identify a translational pathway that represses metabolism to safeguard genome integrity. A system-wide MATRIX survey identified the ancient eIF5A as a pH-regulated translation factor that responds to fermentation-induced acidosis. TMT-pulse-SILAC analysis identified several pH-dependent proteins, including the mTORC1 suppressor Tsc2 and the longevity regulator Sirt1. Sirt1 operates as a pH-sensor that deacetylates nuclear eIF5A during anaerobiosis, enabling the cytoplasmic export of eIF5A/Tsc2 mRNA complexes for translational engagement. Tsc2 induction inhibits mTORC1 to suppress cellular metabolism and prevent acidosis-induced DNA damage. Depletion of eIF5A or Tsc2 leads to metabolic re-initiation and proliferation, but at the expense of incurring substantial DNA damage. We suggest that eIF5A operates as a translatome remodeler that suppresses metabolism to shield the genome.

Modulation of lymphocyte subsets and humoral immune response by florfenicol administered to sheep red blood cell-immunized broiler chickens.

Florfenicol is a broad-spectrum bacteriostatic antibiotic commonly used for the treatment of systemic infections in farm animals. The aim of this study was to determine the effect of florfenicol on the percentage of T lymphocytes (CD3+, CD4+, CD8+, TCRgd+ cells) and B lymphocytes (Bu-1+ cells) and on total serum anti - sheep red blood cell (SRBC) haemagglutinin titer in the peripheral blood of SRBC-immunized broiler chickens. The study included three groups of broiler chickens differentiated by weight (0.5, 1.2, 2.4 kg). Florfenicol was administered orally at a dose of 30 mg/kg. The drug was administered eight times at 24 h intervals. The chickens were immunized with SRBC 24 h after administration of the third dose of florfenicol. Florfenicol increased the percentage of CD3+ blood lymphocytes with a corresponding decrease in the percentage of B lymphocytes in birds weighing 0.5 and 2.4 kg. Florfenicol reduced the production of total anti SRBC-haemagglutinins on day 5 after antigen injection in all three body weight groups of the broiler chickens. In conclusion, florfenicol exerted a modulating effect on the immune response of the birds and this should be taken into consideration when using this antibiotic for certain indications.

Hyperandrogenism and Metabolic Syndrome Are Associated With Changes in Serum-Derived microRNAs in Women With Polycystic Ovary Syndrome.

Polycystic ovary syndrome (PCOS) remains one of the most common endocrine disorder in premenopausal women with an unfavorable metabolic risk profile. Here, we investigate whether biochemical hyperandrogenism, represented by elevated serum free testosterone, resulted in an aberrant circulating microRNA (miRNAs) expression profile and whether miRNAs can identify those PCOS women with metabolic syndrome (MetS). Accordingly, we measured serum levels of miRNAs as well as biochemical markers related to MetS in a case-control study of 42 PCOS patients and 20 Controls. Patients were diagnosed based on the Rotterdam consensus criteria and stratified based on serum free testosterone levels (≥0.034 nmol/l) into either a normoandrogenic (n = 23) or hyperandrogenic (n = 19) PCOS group. Overall, hyperandrogenic PCOS women were more insulin resistant compared to normoandrogenic PCOS women and had a higher prevalence of MetS. A total of 750 different miRNAs were analyzed using TaqMan Low-Density Arrays. Altered levels of seven miRNAs (miR-485-3p, -1290, -21-3p, -139-3p, -361-5p, -572, and -143-3p) were observed in PCOS patients when compared with healthy Controls. Stratification of PCOS women revealed that 20 miRNAs were differentially expressed between the three groups. Elevated serum free testosterone levels, adjusted for age and BMI, were significantly associated with five miRNAs (miR-1290, -20a-5p, -139-3p, -433-3p, and -361-5p). Using binary logistic regression and receiver operating characteristic curves (ROC), a combination panel of three miRNAs (miR-361-5p, -1225-3p, and -34-3p) could correctly identify all of the MetS cases within the PCOS group. This study is the first to report comprehensive miRNA profiling in different subgroups of PCOS women with respect to MetS and suggests that circulating miRNAs might be useful as diagnostic biomarkers of MetS for a different subset of PCOS.

Cell-Enriched Fat Grafting Improves Graft Retention in a Porcine Model: A Dose-Response Study of Adipose-Derived Stem Cells versus Stromal Vascular Fraction.

BACKGROUND: Cell-enrichment of fat grafts has produced encouraging results, but the optimal concentrations and types of added cells are unknown. The authors investigated the effects of enrichment with various concentrations of ex vivo-expanded adipose-derived stem/stromal cells and stromal vascular fraction on graft retention in a porcine model. METHODS: Adipose-derived stem/stromal cells were culture-expanded, and six fat grafts (30 ml) were prepared for each minipig (n = 13). The authors investigated grafts enriched with 2.5 × 10 to 20 × 10 adipose-derived stem cells/ml and stromal vascular fraction and nonenriched control grafts. Each pig served as its own control. Magnetic resonance imaging was performed immediately after grafting and 120 days postoperatively before the pigs were euthanized, and histologic samples were collected. RESULTS: The authors recorded an enhanced relative graft retention rate of 41 percent in a pool of all cell-enriched grafts compared to the nonenriched control (13.0 percent versus 9.2 percent; p = 0.0045). A comparison of all individual groups showed significantly higher graft retention in the 10 × 10-adipose-derived stem/stromal cells per milliliter group compared with the control group (p = 0.022). No significant differences were observed between the cell-enriched groups (p = 0.66). All fat grafts showed a significantly better resemblance to normal fat tissue in the periphery than in the center (p < 0.009), but no differences in overall graft morphology were observed between groups (p > 0.17). CONCLUSIONS: Cell-enriched fat grafting improved graft retention and was feasible in this porcine model. No significant differences in graft retention were observed among the various adipose-derived stem/stromal cell concentrations or between adipose-derived stem/stromal cell and stromal vascular fraction enrichment. Future studies using this model can help improve understanding of the role of adipose-derived stem/stromal cells in cell-enriched fat grafting.

Characterization of Polish Steinernema silvaticum isolates (Nematoda: Steinernematidae) using morphological and molecular data.

Four strains of entomopathogenic nematodes were isolated with a live trap method in southern Poland. The isolates were identified as Steinernema silvaticum based on morphological, morphometric and molecular data. Infective juveniles of Polish S. silvaticum isolates differ in body length from S. weiseri (951 vs 740 µm, respectively), and in the hyaline tail portion from S. kraussei (48 vs 38%, respectively). First-generation males of S. silvaticum are longer than those of S. kraussei, S. weiseri and S. ichnusae (1829 vs 1400, 1180 and 1341 µm, respectively). Males of S. silvaticum and a sister species S. kraussei can be distinguished by the distance from the anterior end to the nerve ring (142 vs 105 µm), spicule (66 vs 49 µm) and gubernaculum length (45 vs 33 µm), and the presence of a mucron. The analysis of internal transcribed spacer (ITS), D2-D3 and cox1 sequences of the tested nematodes revealed differences of 3-5%, 3% and 12-13%, respectively, from S. kraussei strains. The phylogeny of both nuclear and mitochondrial genes indicated close relationships of the Polish S. silvaticum isolates with S. kraussei, S. oregonense and S. cholashanense. The reproductive isolation of the studied isolates was confirmed by hybridization tests with other European feltiae-kraussei group representatives. This study has supplemented the original description of S. silvaticum with morphological and morphometric characterization of the first-generation males and females. This is also the first molecular study of this species based on a multi-gene approach.

A Novel Porcine Model for Future Studies of Cell-enriched Fat Grafting.

BACKGROUND: Cell-enriched fat grafting has shown promising results for improving graft survival, although many questions remain unanswered. A large animal model is crucial for bridging the gap between rodent studies and human trials. We present a step-by-step approach in using the Göttingen minipig as a model for future studies of cell-enriched large volume fat grafting. METHODS: Fat grafting was performed as bolus injections and structural fat grafting. Graft retention was assessed by magnetic resonance imaging after 120 days. The stromal vascular fraction (SVF) was isolated from excised fat and liposuctioned fat from different anatomical sites and analyzed. Porcine adipose-derived stem/stromal cells (ASCs) were cultured in different growth supplements, and population doubling time, maximum cell yield, expression of surface markers, and differentiation potential were investigated. RESULTS: Structural fat grafting in the breast and subcutaneous bolus grafting in the abdomen revealed average graft retention of 53.55% and 15.28%, respectively, which are similar to human reports. Liposuction yielded fewer SVF cells than fat excision, and abdominal fat had the most SVF cells/g fat with SVF yields similar to humans. Additionally, we demonstrated that porcine ASCs can be readily isolated and expanded in culture in allogeneic porcine platelet lysate and fetal bovine serum and that the use of 10% porcine platelet lysate or 20% fetal bovine serum resulted in population doubling time, maximum cell yield, surface marker profile, and trilineage differentiation that were comparable with humans. CONCLUSIONS: The Göttingen minipig is a feasible and cost-effective, large animal model for future translational studies of cell-enriched fat grafting.

Antioxidant defence in the brain of 1-d-old chickens exposed in ovo to acrylamide.

1. Acrylamide (ACR) is a potent neurotoxicant, although information on its toxic influence on the developing neural system is still limited. The effects of in-ovo-injected ACR on the antioxidant system activity in the brain of newly hatched chickens was examined. This model eliminated the mother's contribution to embryonic development. It was also recognised as an adequate model for animal embryonic development. 2. ACR was injected on d 4 of embryogenesis - in doses of 1.25 and 2.50 mg/egg (n = 40 eggs/group/120 eggs). The doses corresponded well with ACR doses used in other animal studies and their concentrations in certain animal feeds. 3. Mortality and incidences of malformations were not found to increase significantly. Significant depletion of glutathione was detected in the cerebellum, cerebrum and medulla oblongata of specimens exposed to the highest doses of ACR. Enzymatic activity was affected by the highest ACR doses. Glutathione peroxidase (GPx) activity increased significantly in the cerebrum, medulla oblongata and the hypothalamus. Superoxide dismutase (SOD) activity increased significantly in hypothalamus and decreased in cerebellum and cerebrum. A significant depletion of catalase (CAT) activity was detected in cerebellum. In the hypothalamus, the increased SOD/GPx and SOD/CAT ratios suggest the risk of H2O2. 4. It was concluded that ACR significantly influences the antioxidative defence in the chicken brain at doses of 1.25 and 2.50 mg/egg.

Standardized assessment of tumor-infiltrating lymphocytes in breast cancer: an evaluation of inter-observer agreement between pathologists.

INTRODUCTION: In breast cancer, there is a growing body of evidence that tumor-infiltrating lymphocytes (TILs) may have clinical utility and may be able to direct clinical decisions for subgroups of patients. Clinical utility is, however, not sufficient for warranting the implementation of a new biomarker in the routine practice, and evaluation of the analytical validity is needed, including testing the reproducibility of decentralized assessment of TILs. The aim of this study was to evaluate the inter-observer agreement of TILs assessment using a standardized method, as proposed by the International TILs Working Group 2014, applied to a cohort of breast cancers reflecting an average breast cancer population. MATERIAL AND METHODS: Stromal TILs were assessed using full slide sections from 124 breast cancers with varying histology, malignancy grade and ER- and HER2 status. TILs were estimated by nine dedicated breast pathologists using scanned hematoxylin-eosin stainings. TILs results were categorized using various cutoffs, and the inter-observer agreement was evaluated using the intraclass coefficient (ICC), Kappa statistics as well as individual overall agreements with the median value of TILs. RESULTS: Evaluation of TILs led to an ICC of 0.71 (95% CI: 0.65-0.77) corresponding to an acceptable agreement. Kappa values were in the range of 0.38-0.46 corresponding to a fair to moderate agreement. The individual agreements increased, when using only two categories ('high' vs. 'low' TILs) and a cutoff of 50-60%. DISCUSSION: The results of the present study are in accordance with previous studies, and shows that the proposed methodology for standardized evaluation of TILs renders an acceptable inter-observer agreement. The findings, however, indicate that assessment of TILs needs further refinement, and is in support of the latest St. Gallen Consensus, that routine reporting of TILs for early breast cancer is not ready for implementation in a clinical setting.

The influence of rapid growth in broilers on florfenicol pharmacokinetics - allometric modelling of the pharmacokinetic and haemodynamic parameters.

1. The aim of this study was to determine if the pharmacokinetics (PK) of florfenicol (FF) undergo age-dependent changes in broilers. Since drug elimination depends on cardiovascular functions, a haemodynamic study was performed in parallel. 2. Broilers of 0.68, 1.27, 2.45 and 5.13 kg were administered FF in a single intravenous dose of 30 mg/kg body weight. Plasma drug concentrations were determined using high-performance liquid chromatography and PK parameters were calculated using a non-compartmental model. Echocardiography was used to measure haemodynamic functions. 3. During growth, the area under the drug concentration-time curve (AUCinf) increased from 25.7 ± 2.9 to 39.0 ± 8.0 mg h/l. Total body clearance (ClB) gradually decreased from 1.19 ± 0.14 to 0.80 ± 0.15 l/h/kg. Elimination half-life increased from 0.73 ± 0.08 to 1.07 ± 0.07 h, whereas volume of distribution (Vss) remained unchanged. Haemodynamic measurements revealed an increase in cardiac output, from 495 ± 65 to 1303 ± 306 ml/min, in the respective body weight groups. 4. Allometric models for PK and haemodynamic parameters were developed and validated. All models proved to be statistically significant; however, only models for ClB and Vss met stringent validation criteria. Model for ClB was used to calculate an optimal dose for a given age group that provides uniform AUCinf. 5. Age-dependent change in FF kinetics may cause variability in therapeutic response under clinical conditions. A novel approach to the dosing protocol was proposed as a means of optimising therapeutic efficacy.

Reoperation Rates in Ductal Carcinoma In Situ vs Invasive Breast Cancer After Wire-Guided Breast-Conserving Surgery.

Importance: New techniques for preoperative localization of nonpalpable breast lesions may decrease the reoperation rate in breast-conserving surgery (BCS) compared with rates after surgery with the standard wire-guided localization. However, a valid reoperation rate for this procedure needs to be established for comparison, as previous studies on this procedure include a variety of malignant and benign breast lesions. Objectives: To determine the reoperation rate after wire-guided BCS in patients with histologically verified nonpalpable invasive breast cancer (IBC) or ductal carcinoma in situ (DCIS) and to examine whether the risk of reoperation is associated with DCIS or histologic type of the IBC. Design, Setting, and Participants: This nationwide study including women with histologically verified IBC or DCIS having wire-guided BCS performed between January 1, 2010, and December 31, 2013, used data from the Danish National Patient Registry that were cross-checked with the Danish Breast Cancer Group database and the Danish Pathology Register. Main Outcomes and Measures: Reoperation rate after wire-guided BCS in patients with IBC or DCIS. Results: Wire-guided BCS was performed in 4118 women (mean [SD] age, 60.9 [8.7] years). A total of 725 patients (17.6%) underwent a reoperation: 593 were reexcisions (14.4%) and 132 were mastectomies (3.2%). Significantly more patients with DCIS (271 of 727 [37.3%]) than with IBC (454 of 3391 [13.4%]) underwent a reoperation (adjusted odds ratio, 3.82; 95% CI, 3.19-4.58; P < .001). After the first reexcision, positive margins were still present in 97 patients (16.4%). The risk of repeated positive margins was significantly higher in patients with DCIS vs those with IBC (unadjusted odds ratio, 2.21; 95% CI, 1.42-3.43; P < .001). The risk of reoperation was significantly increased in patients with lobular carcinoma vs those with ductal carcinoma (adjusted odds ratio, 1.44; 95% CI 1.06-1.95; P = .02). A total of 202 patients (4.9%) had a subsequent completion mastectomy, but no difference was found in the type of reoperation between patients with DCIS and those with IBC. Conclusions and Relevance: A lower reoperation rate after wire-guided BCS was found in this study than those shown in previous studies. However, the risk of reoperation in patients with DCIS was 3 times higher than in those with IBC. The widespread use of mammographic screening will increase the number of patients diagnosed with DCIS, making a precise localization of nonpalpable DCIS lesions even more important.

Overcoming the bottleneck of platelet lysate supply in large-scale clinical expansion of adipose-derived stem cells: A comparison of fresh versus three types of platelet lysates from outdated buffy coat-derived platelet concentrates.

BACKGROUND: Platelet lysates (PL) represent a promising replacement for xenogenic growth supplement for adipose-derived stem cell (ASC) expansions. However, fresh platelets from human blood donors are not clinically feasible for large-scale cell expansion based on their limited supply. Therefore, we tested PLs prepared via three methods from outdated buffy coat-derived platelet concentrates (PCs) to establish an efficient and feasible expansion of ASCs for clinical use. METHODS: PLs were prepared by the freeze-thaw method from freshly drawn platelets or from outdated buffy coat-derived PCs stored in the platelet additive solution, InterSol. Three types of PLs were prepared from outdated PCs with platelets suspended in either (1) InterSol (not manipulated), (2) InterSol + supplemented with plasma or (3) plasma alone (InterSol removed). Using these PLs, we compared ASC population doubling time, cell yield, differentiation potential and cell surface markers. Gene expression profiles were analyzed using microarray assays, and growth factor concentrations in the cell culture medium were measured using enzyme-linked immunosorbent assay (ELISA). RESULTS: Of the three PL compositions produced from outdated PCs, removal of Intersol and resuspension in plasma prior to the first freezing process was overall the best. This specific outdated PL induced ASC growth kinetics, surface markers, plastic adherence and differentiation potentials comparable with PL from fresh platelets. ASCs expanded in PL from fresh versus outdated PCs exhibited different expressions of 17 overlapping genes, of which 10 were involved in cellular proliferation, although not significantly reflected by cell growth. Only minor differences in growth factor turnover were observed. CONCLUSION: PLs from outdated platelets may be an efficient and reliable source of human growth supplement allowing for large-scale ASC expansion for clinical use.

MicroRNAs related to androgen metabolism and polycystic ovary syndrome.

Polycystic ovary syndrome (PCOS) is a frequent endocrine disorder in women. PCOS is associated with altered features of androgen metabolism, increased insulin resistance and impaired fertility. Furthermore, PCOS, being a syndrome diagnosis, is heterogeneous and characterized by polycystic ovaries, chronic anovulation and evidence of hyperandrogenism, as well as being associated with chronic low-grade inflammation and an increased life time risk of type 2 diabetes. A number of androgen species contribute to the symptoms of increased androgen exposure seen in many, though not all, cases of PCOS: Testosterone, androstenedione, dehydroepiandrosterone (DHEA) and dehydroepiandrosterone sulfate (DHEAS), where the quantitatively highest amount of androgen is found as DHEAS. The sulfation of DHEA to DHEAS depends on a number of enzymes, and altered sulfate metabolism may be associated with and contribute to the pathogenesis of PCOS. MicroRNAs (miRNAs) are small, non-coding RNAs that are able to regulate gene expression at the post-transcriptional level. Altered miRNA levels have been associated with diabetes, insulin resistance, inflammation and various cancers. Studies have shown that circulating miRNAs are present in whole blood, serum, plasma and the follicular fluid of PCOS patients and that these might serve as potential biomarkers and a new approach for the diagnosis of PCOS. In this review, recent work on miRNAs with respect to PCOS will be summarized. Our understanding of miRNAs, particularly in relation to PCOS, is currently at a very early stage, and additional studies will yield important insight into the molecular mechanisms behind this complex and heterogenic syndrome.