Wild Type Mesenchymal Cells Contribute to the Lung Pathology of Lymphangioleiomyomatosis

Lymphangioleiomyomatosis (LAM) is a rare disease leading to lungs cysts and progressive respiratory failure. Cells of unknown origin accumulate in the lungs forming nodules and eventually resulting in lung cysts. These LAM cells are described as clonal with bi-allelic mutations in TSC-2 resulting in constitutive mTOR activation. However LAM nodules are heterogeneous structures containing cells of different phenotypes; we investigated whether recruited wild type cells were also present alongside mutation bearing cells. Cells were isolated from LAM lung tissue, cultured and characterised using microscopy, immunocytochemistry and western blotting. Fibroblast-like cells were identified in lung tissue using immunohistochemical markers. Fibroblast chemotaxis toward LAM cells was examined using migration assays and 3D cell culture. Fibroblast-like cells were obtained from LAM lungs: these cells had fibroblast-like morphology, actin stress fibres, full length tuberin protein and suppressible ribosomal protein S6 activity suggesting functional TSC-1/2 protein. Fibroblast Activation Protein, Fibroblast Specific Protein/S100A4 and Fibroblast Surface Protein all stained subsets of cells within LAM nodules from multiple donors. In a mouse model of LAM, tuberin positive host derived cells were also present within lung nodules of xenografted TSC-2 null cells. In vitro, LAM 621-101 cells and fibroblasts formed spontaneous aggregates over three days in 3D co-cultures. Fibroblast chemotaxis was enhanced two fold by LAM 621-101 conditioned medium (p=0.05), which was partially dependent upon LAM cell derived CXCL12. Further, LAM cell conditioned medium also halved fibroblast apoptosis under serum free conditions (p=0.03). Our findings suggest that LAM nodules contain a significant population of fibroblast-like cells. Analogous to cancer associated fibroblasts, these cells may provide a permissive environment for LAM cell growth and contribute to the lung pathology of LAM lung disease.     

From the Schnauzenorgan to the back: Morphological comparison of mormyromast electroreceptor organs at different skin regions of Gnathonemus petersii

The nocturnally active weakly electric fish Gnathonemus petersii is known to employ active electrolocation for the detection of objects and for orientation in its environment. The fish emits pulse‐type electric signals with an electric organ and perceives these signals with more than 3,000 epidermal electroreceptor organs, the mormyromasts, which are distributed over the animal's skin surface. In this study, we measured the metric dimensions of the mormyromasts from different body regions to find structural and functional specialization of the various body parts. We focused on the two foveal regions of G. petersii, which are located at the elongated and movable chin (the Schnauzenorgan; SO) and at the nasal region (NR), the skin region between the mouth and the nares. These two foveal regions were compared to the dorsal part (back) of the fish, which contains typical nonfoveal mormyromasts. While the gross anatomy of the mormyromasts from all skin regions is similar, the metric dimensions of the main substructures differed. The mormyromasts at the SO are the smallest and contain the smallest receptor cells. In addition, the number of receptor cells per organ is lowest at the SO. In contrast, at the back the biggest receptor organs with the highest amount of receptor cells per organ occur. The mormyromasts at the NR are in several respects intermediate between those from the back and the SO. However, mormyromasts at the NR are longer than those at all other skin regions, the canal leading from the receptor pore to the inner chambers were the longest and the overlaying epidermal layers are the thickest. These results show that mormyromasts and the epidermis they are embedded in at both foveal regions differ specifically from those found on the rest of the body. The morphological specializations lead to functional specialization of the two foveae. J. Morphol., 2012. © 2012 Wiley Periodicals, Inc.     

A Dual Role for Corneal Dendritic Cells in Herpes Simplex Keratitis: Local Suppression of Corneal Damage and Promotion of Systemic Viral Dissemination

The cornea is the shield to the foreign world and thus, a primary site for peripheral infections. However, transparency and vision are incompatible with inflammation and scarring that may result from infections. Thus, the cornea is required to perform a delicate balance between fighting infections and preserving vision. To date, little is known about the specific role of antigen-presenting cells in viral keratitis. In this study, utilizing an established murine model of primary acute herpes simplex virus (HSV)-1 keratitis, we demonstrate that primary HSV keratitis results in increased conventional dendritic cells (cDCs) and macrophages within 24 hours after infection. Local depletion of cDCs in CD11c-DTR mice by subconjuntival diphtheria toxin injections, led to increased viral proliferation, and influx of inflammatory cells, resulting in increased scarring and clinical keratitis. In addition, while HSV infection resulted in significant corneal nerve destruction, local depletion of cDCs resulted in a much more severe loss of corneal nerves. Further, local cDC depletion resulted in decreased corneal nerve infection, and subsequently decreased and delayed systemic viral transmission in the trigeminal ganglion and draining lymph node, resulting in decreased mortality of mice. In contrast, sham depletion or depletion of macrophages through local injection of clodronate liposomes had neither a significant impact on the cornea, nor an effect on systemic viral transmission. In conclusion, we demonstrate that corneal cDCs may play a primary role in local corneal defense during viral keratitis and preserve vision, at the cost of inducing systemic viral dissemination, leading to increased mortality.     

Effect of vitamin D3 on behavioural and biochemical parameters in diabetes type 1‐induced rats

Diabetes is associated with long‐term complications in the brain and reduced cognitive ability. Vitamin D₃ (VD₃) appears to be involved in the amelioration of hyperglycaemia in streptozotocin (STZ)‐induced diabetic rats. Our aim was to analyse the potential of VD₃ in avoiding brain damage through evaluation of acetylcholinesterase (AChE), Na⁺K⁺‐adenosine triphosphatase (ATPase) and delta aminolevulinate dehydratase (δ‐ALA‐D) activities and thiobarbituric acid reactive substance (TBARS) levels from cerebral cortex, as well as memory in STZ‐induced diabetic rats. Animals were divided into eight groups (n = 5): control/saline, control/metformin (Metf), control/VD₃, control/Metf + VD₃, diabetic/saline, diabetic/Metf, diabetic/VD₃ and diabetic/Metf + VD₃. Thirty days after treatment, animals were submitted to contextual fear‐conditioning and open‐field behavioural tests, after which they were sacrificed and the cerebral cortex was dissected. Our results demonstrate a significant memory deficit, an increase in AChE activity and TBARS levels and a decrease in δ‐ALA‐D and Na⁺K⁺‐ATPase activities in diabetic rats when compared with the controls. Treatment of diabetic rats with Metf and VD₃ prevented the increase in AChE activity when compared with the diabetic/saline group. In treated diabetic rats, the decrease in Na⁺K⁺‐ATPase was reverted when compared with non‐treated rats, but the increase in δ‐ALA‐D activity was not. VD₃ prevented diabetes‐induced TBARS level and improved memory. Our results show that VD₃ can avoid cognitive deficit through prevention of changes in important enzymes such as Na⁺K⁺‐ATPase and AChE in cerebral cortex in type 1 diabetic rats. Copyright © 2014 John Wiley & Sons, Ltd.     

Molecular recognition and targeting of lysosomal proteins.

Recent studies have established that in mammalian cells insulin-like growth factor-II can couple the large mannose-6-phosphate receptor to a GTP-binding protein and that the insulin-like growth factor-II-induced activation of the GTP-binding protein is inhibited by mannose-6-phosphate and lysosomal enzymes. In mouse, the gene for the large mannose-6-phosphate receptor is maternally imprinted.     

Analysis of the binding of p-chlorophenyl-methoxybenzyl-ketoxime (CPMB-oxime) to mitochondrial cytochrome c reductase.

Cytochrome c reductase is inhibited by p-chlorophenyl-methoxybenzyl-ketoxime (CPMB-oxime). CPMB-oxime induces a red-shift of the reduced spectrum of cytochrome b. The inhibitor blocks the oxidation of ubihydroquinone at the QP center of this enzyme in a non-competitive way. The binding stoichiometry equals one inhibitor molecule per Qp center. The apparent Kd in a red-shift assay was 6.9 +/- 0.6 microM. All binding characteristics analysed in this study were very similar to those of the E-beta-methoxyacrylate inhibitors, although the chemical structure is different from these inhibitors. This result is interpreted as a support for the inhibitory mechanism based on the model of a 'catalytic switch' proposed recently for the E-beta-methoxyacrylate inhibitors (MOA-inhibitors (Brandt and von Jagow, Eur. J. Biochem.     

Meloidogyne incognita Surface Antigen Epitopes in Infected Arabidopsis Roots

Surface-coat epitopes of Meloidogyne incognita were detected in root tissues of Arabidopsis thaliana during migration and feeding site formation. A whole-mount root technique was used for immunolocalization of surface coat epitopes in A. thaliana, with the aid of a monoclonal antibody raised specifically against the outer surface of infective juveniles of M. incognita. The antibody, which was Meloidogyne-specific, recognized a fucosyl-bearing glycoprotein in the surface coat. During migration in host tissues the surface coat was shed, initially accumulating in the intercellular spaces next to the juvenile and later at cell junctions farther from the nematode. Upon induction of giant cell formation, the antibody bound to proximally located companion cells and sieve elements of the phloem.