Малий бізнес в Україні: окремі аспекти сучасного стану та проблеми подальшого розвитку

У статті розглянуто деякі аспекти сучасного стану підприємництва в Україні у порівнянні з розвинутими країнами світу; визначені актуальні проблеми подальшого розвитку малого бізнесу, його роль і потенційні можливості у подоланні кризових явищ в економіці держави; окреслена необхідність істотного покращення державної регуляторної політики у сфері підприємництва, нагальні потреби підвищення якості управління бізнесовими структурами.In the article main aspects of current state of entrepreneurship in Ukraine are reviewed in comparison with developed countries, problems of small and medium business further development and it’s role and potential ability in overcoming crisis event in country economy are determined, necessity of substantial improvement of government regulation politic and also emergency need of business structures management quality improvement are presented

ESR1 Amplification in Breast Cancer by Optimized RNase FISH: Frequent but Low-Level and Heterogeneous. PLoS ONE 8(12): e84189. Optical Dataset S1

<p><strong>Moelans CB, Holst F, Hellwinkel O, Simon R, van Diest PJ (2013) <em>ESR1</em> Amplification in Breast Cancer by Optimized RNase FISH: Frequent but Low-Level and Heterogeneous. PLoS ONE 8(12): e84189. doi:10.1371/journal.pone.0084189</strong></p> <p><strong>Optical Dataset S1 (without RNase) of S1+S2</strong></p> <p>OD S2: http://dx.doi.org/10.6084/m9.figshare.864525</p> <p><strong>Combined Description for Optical Dataset S1 and S2:</strong></p> <p>The optical datasets illustrate the FISH signal appearance by full size <em>ESR1</em> FISH photographs (not all of them representative) of a total of 35 breast cancer cases (Case#1-35) including 24 tumors with increased <em>ESR1</em> copy numbers by FISH (15 <em>ESR1</em> amplified and 9 <em>ESR1</em> gained, Case#01-#24) and 11 tumors without <em>ESR1</em> copy number increase (Case#25-#35), selected from a consecutive subset of 90 formalin fixed, paraffin embedded grade 3 breast cancers from the archives of the Department of Pathology in Hamburg and previously used in an <em>ESR1</em> FISH mapping study [12]. To illustrate the results of RNase A treatment, for all 24 cases with <em>ESR1</em> copy number increase different pictures (Fig.01-19) with (+RNase A: optical dataset S2; “OD2”) and without RNase A treatment (optical dataset S1; “OD1”) are available.</p> <p>FISH data are optical patterns visualized by imaging technology and translated by subjective interpretation into numerical information. To support the confirmability of optical FISH “raw data”, the pictures document by three color photographs the <em>ESR1</em> FISH signal appearance in nuclei with <em>ESR1</em> copy number increase and nuclei without <em>ESR1</em> copy number increase on 4µm conventional large section FFPE tissue slides, showing <em>ESR1</em> signals (green), <em>CEN6</em> signals (orange) and cell nuclei (blue) at 100x or 63x magnification. Pictures are labeled “CNI” for observed gene “copy number increase” and “NO” for “normal” or “no copy number increase”. Gene loci with additional allelic copies (CNI) are marked exemplarily in some cases (white arrows and edges), partially indicating occurrence of nuclei with “CNI” appearing sporadically distributed in a pattern of mosaic heterogeneity intermingled with nuclei without “CNI”.</p> <p>Due to intensity variations, nucleus truncation during slide sectioning and the three dimensional distribution of signals, individual gene copies may not always become visible. To illustrate the distribution of signals within the z-axis, some pictures are taken with different z-layers (Z-Stack A-D).</p> <p>To illustrate the difference between cases or tissue areas showing <em>ESR1</em> copy number increase (CNI) and cases or tissue areas without <em>ESR1</em> copy number increase, pictures of the 11 cases and other tissue areas with clearly normal copy number or “no copy number increase” (NO) are shown in addition.</p> <p>12. Holst F, Stahl PR, Ruiz C, Hellwinkel O, Jehan Z, et al. (2007) Estrogen receptor alpha (ESR1) gene amplification is frequent in breast cancer. Nat Genet 39: 655-660.</p

Effect of RNase treatment on FISH and on correlation between FISH and MLPA.

<p>(a) RNase pretreatment resulted in a higher fraction of tumor cells showing point-shaped FISH signals, by eliminating eye catching fuzzy clusters of ESR1 signals seen by standard FISH (b) MLPA copy number ratios in FISH “not increased”, gained and amplified samples without and with RNase treatment.</p

ESR1 Amplification in Breast Cancer by Optimized RNase FISH: Frequent but Low-Level and Heterogeneous. PLoS ONE 8(12): e84189. Optical Dataset S2

<p><strong>Moelans CB, Holst F, Hellwinkel O, Simon R, van Diest PJ (2013) <em>ESR1</em> Amplification in Breast Cancer by Optimized RNase FISH: Frequent but Low-Level and Heterogeneous. PLoS ONE 8(12): e84189. doi:10.1371/journal.pone.0084189</strong></p> <p><strong>Optical Dataset S2 (with RNase) of S1+S2</strong></p> <p>OD S1: http://dx.doi.org/10.6084/m9.figshare.864194</p> <p><strong>Combined Description for Optical Dataset S1 and S2:</strong></p> <p>The optical datasets illustrate the FISH signal appearance by full size <em>ESR1</em> FISH photographs (not all of them representative) of a total of 35 breast cancer cases (Case#1-35) including 24 tumors with increased <em>ESR1</em> copy numbers by FISH (15 <em>ESR1</em> amplified and 9 <em>ESR1</em> gained, Case#01-#24) and 11 tumors without <em>ESR1</em> copy number increase (Case#25-#35), selected from a consecutive subset of 90 formalin fixed, paraffin embedded grade 3 breast cancers from the archives of the Department of Pathology in Hamburg and previously used in an <em>ESR1</em> FISH mapping study [12]. To illustrate the results of RNase A treatment, for all 24 cases with <em>ESR1</em> copy number increase different pictures (Fig.01-19) with (+RNase A: optical dataset S2; “OD2”) and without RNase A treatment (optical dataset S1; “OD1”) are available.</p> <p>FISH data are optical patterns visualized by imaging technology and translated by subjective interpretation into numerical information. To support the confirmability of optical FISH “raw data”, the pictures document by three color photographs the <em>ESR1</em> FISH signal appearance in nuclei with <em>ESR1</em> copy number increase and nuclei without <em>ESR1</em> copy number increase on 4µm conventional large section FFPE tissue slides, showing <em>ESR1</em> signals (green), <em>CEN6</em> signals (orange) and cell nuclei (blue) at 100x or 63x magnification. Pictures are labeled “CNI” for observed gene “copy number increase” and “NO” for “normal” or “no copy number increase”. Gene loci with additional allelic copies (CNI) are marked exemplarily in some cases (white arrows and edges), partially indicating occurrence of nuclei with “CNI” appearing sporadically distributed in a pattern of mosaic heterogeneity intermingled with nuclei without “CNI”.</p> <p>Due to intensity variations, nucleus truncation during slide sectioning and the three dimensional distribution of signals, individual gene copies may not always become visible. To illustrate the distribution of signals within the z-axis, some pictures are taken with different z-layers (Z-Stack A-D).</p> <p>To illustrate the difference between cases or tissue areas showing <em>ESR1</em> copy number increase (CNI) and cases or tissue areas without <em>ESR1</em> copy number increase, pictures of the 11 cases and other tissue areas with clearly normal copy number or “no copy number increase” (NO) are shown in addition.</p> <p>12. Holst F, Stahl PR, Ruiz C, Hellwinkel O, Jehan Z, et al. (2007) Estrogen receptor alpha (ESR1) gene amplification is frequent in breast cancer. Nat Genet 39: 655-660.</p

Clinicopathological correlation of BLCAP expression in 2,197 sample TMA.

*<p>Kruskal-Wallis one way analysis of variance on ranks.</p>**<p>Mann-Whitney rank-sum test.</p

Differential expression of BLCAP in tumor samples.

<p>The DCTB 123 patient set, including normal, tumor and lymph node metastasis samples (A), or a subset of matched 62 normal and tumor samples (B) were analyzed by quantitative IHC. Mean intensity scores for each group are indicated by red lines.</p

Immunohistochemical expression analysis of BLCAP in FFPE breast tissue samples.

<p>(A) No immunostaining was observed in a normal breast tissue section reacted with BLCAP antibody preincubated with immunizing peptide. (B) Immunohistochemical staining of BLCAP protein in a normal breast tissue sample demonstrated the presence of the BLCAP antigen in luminal epithelial cells with weak cytoplasmic expression (black arrow). Marked nuclear expression was also observed occasionally (red arrow). Yellow arrow points to a vessel with moderate immunoreactivity for BLCAP. (C) In a few cases IHC analysis of tumor samples showed that BLCAP was expressed in tumor cells with weak cytoplasmic expression (red arrow). (D) Most cases showed moderate to strong cytoplasmic expression with no detectable nuclear presence (red arrow) but in some cases (E) we could observe strong nuclear expression of BLCAP (red arrow). (F) In a few cases, samples were heterogenous with some cells showing distinct perinuclear immunoreactivity for BLCAP (red arrow). (G) Malignant cells showed stronger immunoreactivity (red arrows) than adjacent normal-looking ducts (black arrow), demonstrating up-regulation of this protein in tumor cells. Yellow arrows point to vessels with strong immunoreactivity for BLCAP. (H) We also observed up-regulation of BLCAP in early lesions where lobular carcinoma in situ cells showed overexpression of this protein (grey arrows) in relation to normal adjacent areas (black arrow), and at levels comparable to invasive carcinoma cells (red arrow).</p

Clinical characteristics of DCTB tissue specimens.

<p>Clinical characteristics of DCTB tissue specimens.</p

2D PAGE and 2D Western blot analysis of BLCAP protein spot patterns.

<p>(A) COS-1 cells transfected with pZeoSV2 empty vector and labeled with ³⁵S-methionine. (B) COS-1 cells transfected with pZeoSV2– BLCAP overexpressing construct and labeled with ³⁵S-methionine. Radioactive metabolic labeling (³⁵S-methionine) of COS-1 cells was used to ensure the highest detection sensitivity. (C) 2D Western blot of COS-1 cells transfected with pZeoSV2– BLCAP construct detected with anti-BLCAP antibody (10 sec film exposure). (D) 2D gel of proteins from breast tumor 63 stained with silver. (E) 2D Western blot of protein lysate from breast tumor 63 (see D) reacted with anti-BLCAP antibody (1 min film exposure). The positions of the BLCAP protein in the 2D-PAGE gels and corresponding 2D Western blots, are indicated by black arrows. The positions of several reference proteins are indicated by red arrows: ACTB – beta actin; ENO1 -alpha enolase 1; CANX – calnexin; PDI - Protein disulfide-isomerase; TUBA1A - tubulin alpha-1A chain; YWHAZ - 14-3-3 protein zeta/delta. The identity of all reference spots were confirmed by MS analysis.</p

Expression analysis of BLCAP by quantitative IHC of normal specimens (blue bars) and corresponding tumor samples (red bars) of 62 matched cases from the DCTB dataset.

<p>Illustrative IHC images are shown for normal and tumor samples with weak immunoreactivity (N11 and T76, respectively), and for normal and tumor samples showing substantial immunoreactivity for BLCAP (N14 and T63, respectively). Magnification 20X.</p