Antibody-functionalized fluid-permeable surfaces for rolling cell capture at high flow rates

Adhesion-based cell capture on surfaces in microfluidic devices forms the basis of numerous biomedical diagnostics and in vitro assays. However, the performance of these platforms is partly limited by interfacial phenomena that occur at low Reynolds numbers. In contrast, cell homing to porous vasculature is highly effective in vivo during inflammation, stem cell trafficking, and cancer metastasis. Here, we show that a porous, fluid-permeable surface functionalized with cell-specific antibodies promotes efficient and selective cell capture in vitro. This architecture is advantageous due to enhanced transport as streamlines are diverted toward the surface. Moreover, specific cell-surface interactions are promoted due to reduced shear, allowing gentle cell rolling and arrest. Together, these synergistic effects enable highly effective cell capture at flow rates more than an order of magnitude larger than those provided by existing devices with solid surfaces.     

Association of Toll-like receptor (TLR) 2, 3 and 9 genes polymorphism with prostate cancer risk in North Indian population

Prostate cancer (PCa) is the most common cancer among men. It has been suggested that toll like receptors (TLRs) may contribute to PCa pathogenesis by stimulating prostate epithelial cell proliferation in response to infectious stimuli. We performed case control study to analyze the genetic variants of TLR2, 3 and 9 gene polymorphisms with PCa risk in a North Indian population. For this study we genotyped age matched, unrelated 195 PCa patients and 250 healthy controls of similar ethnicity in a case-control study. They were genotyped for TLR2 (-196 to -174 Del), TLR3 (c.1377C/T) [rs3775290] and TLR9 (G2848A) [rs352140] gene polymorphisms using polymerase chain reaction and restriction fragment length polymorphism method. Variant allele Del (D) carriers i.e. (ID + DD) of TLR2 (-196 to -174 Del) SNP, demonstrated 1.57 fold increased risk (p = 0.040; OR = 1.57, 95% CI = 1.02-2.24) as compared to Ins (I) allele, suggesting a dominant effect model involved in the risk of this polymorphism in PCa. However, variants of TLR3 and 9 gene polymorphisms were not associated with PCa risk. Our results suggested the low penetrance variant of TLR2 (-196 to -174 Del) to be at increased PCa risk in North Indian population. Functional studies in ethnically diverse populations may provide a more comprehensive involvement of innate immunity in identifying the disease-associated variants for PCa etiology.     

Smoothing of the GB1 Hairpin Folding Landscape by Interfacial Confinement

We study the effects of confinement between planar walls on the folding thermodynamics of a β-hairpin, using large-scale replica-exchange molecular-dynamics simulations with an all-atom model and explicit solvent. We find that the folding free-energy landscape of this peptide observed in bulk is significantly modified when the peptide is confined between the walls. Most notably, the propensity of the peptide to form a misfolded state observed in the bulk solution becomes negligible under confinement. The absence of the misfolded state under confinement can be explained by an increased tendency of hydrophobic aromatic side chains to stay near the walls, because the misfolded state is characterized by a nonnative arrangement of aromatic side chains. These results from a simple confinement model may provide clues about the role of chaperonin confinement in smoothing folding landscapes by avoiding trapped intermediates.     

Response of the chromatophores in relation to the healing of skin wounds in an Indian Major Carp, Labeo rohita (Hamilton)

Chromatophores show significant changes during healing of skin wounds in Labeo rohita (Common Name - Rohu). Wound area can be divided into regions I, II and III. After infliction of wound, skin colour becomes significantly dark by 2 h that is gradually restored by 2 d. In regions II and III at 5 min, epidermal melanophores appear with beaded dendrites. In these regions at 2 h and in region I at 6 h, epidermal melanophores appear small, rounded or irregular shaped having dendritic processes with aggregated melanosomes. Subsequently, melanophores appear having elongated dendrites with dispersed or aggregated melanosomes. At 24 h, clusters of pigmented bodies appear in regions I and II. These bodies increase up to 2 d, and then diminish gradually and disappear by 8 d. Changes in dermal melanophores in region II at 5 min indicate the onset of degeneration. Degenerating melanophores increase up to 12 h, then gradually decline, and disappear by 4 d. Simultaneously, stellate melanophore reappear, gradually increase and appear like control by 8 d. Dermal melanophores in region III at different intervals appear stellate. In region I stellate dermal melanophores appear at 4 d. Stellate melanophores in all regions show different distribution of dispersed or aggregated melanosomes. With the appearance of dermal melanophores, highly refractive, crystalline structures, possibly the refractive platelets of the iridophores, are visualized around them. At subsequent intervals, these are frequently observed. This study provides interesting insights in injury induced changes in chromatophores in fish. The findings could be considered useful in perception of intriguing features in the development of pigment research in future.     

Significant role of estrogen and progesterone receptor sequence variants in gallbladder cancer predisposition: a multi-analytical strategy

Background

Carcinoma of gallbladder (GBC) is an aggressive malignancy. The higher incidence of gallbladder cancer in women has been partly attributed to hormonal factors. Therefore the present study was designed to explore the role of genetic variants in estrogen (ESR1, ESR2) and progesterone (PGR) receptors in conferring risk of gallbladder cancer.

Materials and Methods

The present case-control study recruited total of 860 subjects, including 410 GBC patients, 230 gallstone patients and 220 controls. We examined the associations of 6 selected polymorphisms in three genes: ESR1 (rs2234693, rs9340799, rs1801132), ESR2 (rs1271572, rs1256049) and PGR (rs1042838) with GBC risk. Genotyping for all the polymorphisms was done using PCR-RFLP. Multifactor dimensionality reduction and classification and regression tree approaches were combined with logistic regression to discover high-order gene-gene interactions in hormonal pathway.

Results

On comparing the genotype frequency distribution in gallstone and GBC patients with that of healthy subjects, the homozygous variant genotypes of ESR1-397TT (rs2234693) polymorphism showed significant risk for developing gallstone [odds ratio: OR = 2.9] and GBC [OR = 1.8] respectively. Detailed haplotypes analysis suggested that ESR1 T _rs2234693G _rs9340799C _rs1801132 have significant association in conferring risk for both gallstones [OR = 2.2] and GBC [OR = 3.0]. However, the variant-containing genotypes (DI+II) of PGR (rs1042838) showed low risk in both GBC [OR = 0.4] and gallstone patients [OR = 0.4].On performing the MDR analysis, ESR1 IVS1-397C>T, ESR1 IVS1-351A>G, and ESR2-789 A>C yielded the highest testing accuracy of 0.634. These results were further supported by the CART analysis which revealed that individuals with the combined genotypes of ESR1-397 CT or TT, ESR1-351 AG or GG and ESR2 -789 AA had the highest risk for GBC [OR  = 3.9].

Conclusion

Using multi-analytical approaches, our study showed important role of ESR1 IVS1-397C>T, ESR1 IVS1-351A>G, and ESR2-789 A>C variants in GBC susceptibility and the risk appears to be mediated through gallstone dependent pathway.     

Utility of MLPA in mutation analysis and carrier detection for Duchenne muscular dystrophy

CONTEXT:

Multiplex ligation probe amplification (MLPA) is a new technique to identify deletions and duplications and can evaluate all 79 exons in dystrophin gene in patients with Duchenne muscular dystrophy (DMD). Being semi-quantitative, MLPA is also effective in detecting duplications and carrier testing of females; both of which cannot be done using multiplex PCR. It has found applications in diagnostics of many genetic disorders.

AIM:

To study the utility of MLPA in diagnosis and carrier detection for DMD.

MATERIALS AND METHODS:

Mutation analysis and carrier detection was done by multiplex PCR and MLPA and the results were compared.

RESULTS AND CONCLUSIONS:

We present data showing utility of MLPA in identifying mutations in cases with DMD/BMD. In the present study using MLPA, we identified mutations in additional 5.6% cases of DMD in whom multiplex PCR was not able to detect intragenic deletions. In addition, MLPA also correctly confirmed carrier status of two obligate carriers and revealed carrier status in 6 of 8 mothers of sporadic cases.     

Gene expression profiling demonstrates a novel role for foetal fibrocytes and the umbilical vessels in human fetoplacental development

There is a difference in the susceptibility to inflammation between the umbilical vein (UV) and the umbilical arteries (UAs). This led us to hypothesize that there is an intrinsic difference in the pro-inflammatory response between UA and UV. Real-time quantitative RT-PCR and microarray analysis revealed higher expression of interleukin (IL)-1β and IL-8 mRNA in the UV and differential expression of 567 genes between the UA and UV associated with distinct biological processes, including the immune response. Differential expression of human leukocyte antigen (HLA)-DRA mRNA between the UA and UV was due to unexpected HLA-DR⁺ cells migrating via the umbilical vessels into Wharton's jelly, more frequently in the UV. A significant proportion of these cells co-expressed CD45 and type I pro-collagen, and acquired CD163 or α-smooth muscle actin immunoreactivity in Wharton's jelly. Migrating cells were also found in the chorionic and stem villous vessels. Furthermore, the extent of migration increased with progression of gestation, but diminished in intrauterine growth restriction (IUGR). The observations herein strongly suggest that circulating foetal fibrocytes, routing via umbilical and placental vessels, are a reservoir for key cellular subsets in the placenta. This study reports fibrocytes in the human umbilical cord and placenta for the first time, and a novel role for both circulating foetal cells and the umbilical vessels in placental development, which is deranged in IUGR.