Automatic image segmentation of nuclear stained breast tissue sections using color active contour model and an improved watershed method

Automatic image segmentation of immunohistologically stained breast tissue sections helps pathologists to discover the cancer disease earlier. The detection of the real number of cancer nuclei in the image is a very tedious and time consuming task. Segmentation of cancer nuclei, especially touching nuclei, presents many difficulties to separate them by traditional segmentation algorithms. This paper presents a new automatic scheme to perform both classification of breast stained nuclei and segmentation of touching nuclei in order to get the total number of cancer nuclei in each class. Firstly, a modified geometric active contour model is used for multiple contour detection of positive and negative nuclear staining in the microscopic image. Secondly, a touching nuclei method based on watershed algorithm and concave vertex graph is proposed to perform accurate quantification of the different stains. Finally, benign nuclei are identified by their morphological features and they are removed automatically from the segmented image for positive cancer nuclei assessment. The proposed classification and segmentation schemes are tested on two datasets of breast cancer cell images containing different level of malignancy. The experimental results show the superiority of the proposed methods when compared with other existing classification and segmentation methods. On the complete image database, the segmentation accuracy in term of cancer nuclei number is over than 97%, reaching an improvement of 3–4% over earlier methods.     

PCR-Based Screening for the Most Prevalent Alpha 1 Antitrypsin Deficiency Mutations (PI S, Z, and Mmalton) in COPD Patients from Eastern Tunisia

It is generally agreed that the protease inhibitor (PI) alleles PI*S (Val264Glu) and PI*Z (Lys342Glu) are the most common alpha 1 antitrypsin deficiency variants worldwide, but the PI*Mmalton allele (ΔPhe52) prevails over these variants in some Mediterranean regions. In eastern Tunisia (Mahdia), we screened 100 subjects with chronic obstructive pulmonary disease for these variants. The PI*S and PI*Z alleles were genotyped by the previously described SexAI/Hpγ99I RFLP–PCR. We provide here a new method for PI*Mmalton genotyping using mismatched RFLP–PCR. These methods are suitable for routine clinical application and can easily be reproduced by several laboratories, since they do not require extensive optimization, unlike the previously described bidirectional allele-specific amplification PCR for PI*Mmalton genotyping. Our results were in agreement with previous reports from central Tunisia (Kairouan), suggesting that the PI*Mmalton mutation is the most frequent alpha 1 antitrypsin deficiency-related mutation in Tunisia.     

Novel splicing dysferlin mutation causing myopathy with intra-familial heterogeneity

Molecular Biology Reports - Dysferlinopathies belong to the heterogeneous group of autosomal recessive muscular disorders, caused by mutations in the dysferlin gene and characterized by a high...     

TGF-β1 induces EMT reprogramming of porcine bladder urothelial cells into collagen producing fibroblasts-like cells in a Smad2/Smad3-dependent manner

Activation of fibroblasts and their differentiation into myofibroblasts, excessive collagen production and fibrosis occurs in a number of bladder diseases. Similarly, conversion of epithelial cells into mesenchymal cells (EMT) has been shown to increase fibroblasts like cells. TGF-β1 can induce the EMT and the role of TGF-β1-induced EMT during bladder injury leading to fibrosis and possible organ failure is gaining increasing interest. Here we show that EMT and fibrosis in porcine bladder urothelial (UC) cells are Smad dependent. Fresh normal porcine bladder urothelial cells were grown in culture with or without TGF-β1 and EMT markers were assessed. TGF-β1 treatment induced changes in cellular morphology as depicted by a significant decrease in the expression of E-cadherin and corresponding increase in N-cadherin and α-SMA. We knocked down Smad2 and Smad3 by Smad specific siRNA. Downregulation of E-cadherin expression by TGF-β1 was Smad3-dependent, whereas N-cadherin and α-SMA were dependent on both Smad2 and Smad3. Connective tissue growth factor (CTGF/CCN2), matrix metalloproteinase-2 and -9 (MMP-2, MMP-9) has been shown to play important roles in the pathogenesis of fibrosis. Induction of these genes by TGF-β1 was found to be time dependent. Upregulation of CTGF/CCN2 by TGF-β1 was Smad3 dependent; whereas MMP-2 was Smad2 dependent. Smad2 and Smad3 both participated in MMP-9 expression. TGF-β1 reprogrammed mesenchymal fibroblast like cells robustly expressed collagen I and III and these was inhibited by SB-431542, a TGF-β receptor inhibitor. Our results indicate that EMT of porcine bladder UC cells is TGF-β1 dependent and is mediated through Smad2 and Smad3. TGF-β1 may be an important factor in the development of bladder fibrosis via an EMT mechanism. This identifies a potential amenable therapeutic target.     

Enhanced Expression of <Emphasis Type="Italic">AtNHX1</Emphasis>, in Transgenic Groundnut (<Emphasis Type="Italic">Arachis hypogaea</Emphasis> L.) Improves Salt and Drought Tolerence

Salinity and drought are main threat to agriculture productivity, to avoid further losses it is necessary to improve the genetic material of crops against these stresses In this present study, AtNHX1, a vacuolar type Na⁺/H⁺ antiporter gene driven by 35S promoter was introduced into groundnut using Agrobacterium tumefaciens transformation system. The stable integration of the AtNHX1 gene was confirmed by polymerase chain reaction (PCR) and southern blot analysis. It was found that transgenic plants having AtNHX1 gene are more resistant to high concentration of salt and water deprivation than the wild type plants. Salt and proline level in the leaves of the transgenic plants were also much higher than that of wild type plants. The results showed that overexpression of AtNHX1 gene not only improved salt tolerance but also drought tolerance in transgenic groundnut. Our results suggest that these plants could be cultivated in salt and drought-affected soils.     

Variation in volatile leaf oils of five Eucalyptus species harvested from Jbel Abderrahman arboreta (Tunisia)

Hydrodistillation of the dried leaves of five species of the genus Eucalyptus L' Hér ., viz., E. dundasii Maiden , E. globulus Labill ., E. kitsoniana Maiden , E. leucoxylon F. Muell ., and E. populifolia Hook ., harvested from Jbel Abderrahman arboreta (region of Nabeul, northeast of Tunisia) in April 2006, afforded essential oils in yields varying from 0.9±0.3 to 3.8±0.6%, dependent on the species. E. globulus and E. Kitsoniana provided the highest and the lowest percentage of essential oil amongst the species examined, respectively. Analysis by GC (RI) and GC/MS allowed the identification of 127 compounds, representing 93.8 to 98.7% of the total oil composition. The contents of the different samples varied according to the species. The main components were 1,8‐cineole ( 2 ; 4.7–59.2%), followed by α‐pinene ( 1 ; 1.9–23.6%), trans‐pinocarveol ( 6 ; 3.5–21.6%), globulol ( 8 ; 4.3–12.8%), p‐cymene ( 3 ; 0.5–6.7%), α‐terpineol (1.5–4.5%), borneol (0.2–4.4%), pinocarvone (1.1–3.8%), aromadendrene (1.4–3.4%), isospathulenol (0.0–1.9%), fenchol ( 4 ; 0.1–2.5%), limonene (1.0–2.4%), epiglobulol (0.6–2.1%), viridiflorol ( 9 ; 0.8–1.8%), and spathulenol (0.1–1.6%). E. leucoxylon was the richest species in 2 . Principal component analysis (PCA) and hierarchical cluster analysis (HCA) separated the five Eucalyptus leaf essential oils into four groups, each constituting a chemotype.     

The stimulatory Gα(s) protein is involved in olfactory signal transduction in Drosophila

Seven-transmembrane receptors typically mediate olfactory signal transduction by coupling to G-proteins. Although insect odorant receptors have seven transmembrane domains like G-protein coupled receptors, they have an inverted membrane topology, constituting a key difference between the olfactory systems of insects and other animals. While heteromeric insect ORs form ligand-activated non-selective cation channels in recombinant expression systems, the evidence for an involvement of cyclic nucleotides and G-proteins in odor reception is inconsistent. We addressed this question in vivo by analyzing the role of G-proteins in olfactory signaling using electrophysiological recordings. We found that Gα_s plays a crucial role for odorant induced signal transduction in OR83b expressing olfactory sensory neurons, but not in neurons expressing CO₂ responsive proteins GR21a/GR63a. Moreover, signaling of Drosophila ORs involved Gα_s also in a heterologous expression system. In agreement with these observations was the finding that elevated levels of cAMP result in increased firing rates, demonstrating the existence of a cAMP dependent excitatory signaling pathway in the sensory neurons. Together, we provide evidence that Gα_s plays a role in the OR mediated signaling cascade in Drosophila.