Dynamic interactions between growth factors and extracellular matrix (ECM) are integral to wound healing. These interactions take several forms that may be categorized as direct or indirect. The ECM can directly bind to and release certain growth factors (e.g., heparan sulfate binding to fibroblast growth factor-2), which may serve to sequester and protect growth factors from degradation, and/or enhance their activity. Indirect interactions include binding of cells to ECM via integrins, which enables cells to respond to growth factors (e.g., integrin binding is necessary for vascular endothelial growth factor-induced angiogenesis) and can induce growth factor expression (adherence of monocytes to ECM stimulates synthesis of platelet-derived growth factor). Additionally, matrikines, or subcomponents of ECM molecules, can bind to cell surface receptors in the cytokine, chemokine, or growth factor families and stimulate cellular activities (e.g., tenascin-C and laminin bind to epidermal growth factor receptors, which enhances fibroblast migration). Growth factors such as transforming growth factor-β also regulate the ECM by increasing the production of ECM components or enhancing synthesis of matrix degrading enzymes. Thus, the interactions between growth factors and ECM are bidirectional. This review explores these interactions, discusses how they are altered in difficult to heal or chronic wounds, and briefly considers treatment implications.

https://onlinelibrary.wiley.com/doi/pdf/10.1111/j.1524-475X.2009.00466.x

Interactions between extracellular matrix and growth factors in wound healing.

Amniotic membrane (AM) or amnion is a thin membrane on the inner side of the fetal placenta; it completely surrounds the embryo and delimits the amniotic cavity, which is filled by amniotic liquid. In recent years, the structure and function of the amnion have been investigated, particularly the pluripotent properties of AM cells, which are an attractive source for tissue transplantation. AM has anti-inflammatory, anti-bacterial, anti-viral and immunological characteristics, as well as anti-angiogenic and pro-apoptotic features. AM is a promoter of epithelialization and is a non-tumorigenic tissue and its use has no ethical problems. Because of its attractive properties, AM has been applied in several surgical procedures related to ocular surface reconstruction and the genito-urinary tract, skin, head and neck, among others. So far, the best known and most auspicious applications of AM are ocular surface reconstruction, skin applications and tissue engineering. However, AM can also be applied in oncology. In this area, AM can prevent the delivery of nutrients and oxygen to cancer cells and consequently interfere with tumour angiogenesis, growth and metastasis.

Amniotic membrane: from structure and functions to clinical applications

Taste receptor cells are responsible for transducing chemical stimuli from the environment and relaying information to the nervous system. Bitter, sweet and umami stimuli utilize G-protein coupled receptors which activate the phospholipase C (PLC) signaling pathway in Type II taste cells. However, it is not known how these cells communicate with the nervous system. Previous studies have shown that the subset of taste cells that expresses the T2R bitter receptors lack voltage-gated Ca2+ channels, which are normally required for synaptic transmission at conventional synapses. Here we use two lines of transgenic mice expressing green fluorescent protein (GFP) from two taste-specific promoters to examine Ca2+ signaling in subsets of Type II cells: T1R3-GFP mice were used to identify sweet- and umami-sensitive taste cells, while TRPM5-GFP mice were used to identify all cells that utilize the PLC signaling pathway for transduction. Voltage-gated Ca2+ currents were assessed with Ca2+ imaging and whole cell recording, while immunocytochemistry was used to detect expression of SNAP-25, a presynaptic SNARE protein that is associated with conventional synapses in taste cells. Depolarization with high K+ resulted in an increase in intracellular Ca2+ in a small subset of non-GFP labeled cells of both transgenic mouse lines. In contrast, no depolarization-evoked Ca2+ responses were observed in GFP-expressing taste cells of either genotype, but GFP-labeled cells responded to the PLC activator m-3M3FBS, suggesting that these cells were viable. Whole cell recording indicated that the GFP-labeled cells of both genotypes had small voltage-dependent Na+ and K+ currents, but no evidence of Ca2+ currents. A subset of non-GFP labeled taste cells exhibited large voltage-dependent Na+ and K+ currents and a high threshold voltage-gated Ca2+ current. Immunocytochemistry indicated that SNAP-25 was expressed in a separate population of taste cells from those expressing T1R3 or TRPM5. These data indicate that G protein-coupled taste receptors and conventional synaptic signaling mechanisms are expressed in separate populations of taste cells. The taste receptor cells responsible for the transduction of bitter, sweet, and umami stimuli are unlikely to communicate with nerve fibers by using conventional chemical synapses.

/pdf/mouse-taste-cells-with-g-protein-coupled-taste-receptors-4jbpg04wka.pdf

Mouse taste cells with G protein-coupled taste receptors lack voltage-gated calcium channels and SNAP-25

Corneal myofibroblast biology and pathobiology: generation, persistence, and transparency.

Purpose Peripheral and limbal corneal stromal cells (PLCSCs), which contain keratocytes, have a complex phenotype. Knowledge of keratocyte cell properties, function, and origin is limited. Evidence available thus far has suggested both mesenchymal stromal and hematopoietic characteristics. Multipotent mesenchymal stromal cells (MSCs) are found in an increasing number of tissues and are the subject of considerable interest and investigation in the disciplines of tissue engineering, immunology, gene therapy, and oncology. Methods Isolated PLCSCs were characterized by markers aldehyde dehydrogenase and keratocan, cultured, and analyzed against a set of criteria for the identification of MSCs developed by the International Society of Cellular Therapy (ISCT). PLCSCs were directly compared to fetal liver MSCs (flMSCs). Additional cell surface markers were also used to quantify differentiation, which was also performed on both cell types. Results PLCSCs were found to be plastic adherent, displayed the correct profile and proportions of CSMs, and demonstrated trilineage potential in accordance with the ISCT guidelines. Furthermore, PLCSCs displayed a high degree of similarity to flMSCs and this likeness extended into the non-ISCT MSC cell surface markers and trilineage differentiation, which were often but not always comparable. Conclusions Herein we report a novel observation that PLCSCs conform to all the ISCT criteria and are therefore MSCs. Furthermore, this study has identified the limbal stroma as yet another MSC niche and presents a new perspective on the role of the PLCSC.

Mesenchymal stem cells in the human corneal limbal stroma.

PURPOSE. The gene encoding transcription factor Pax6 resides at the top of a genetic hierarchy controlling development and morphogenesis of the eye. Pax6 continues to be expressed in the ocular surface epithelia of the postnatal eye. The goal of this study was to investigate a possible role for Pax6 in controlling dynamics of the ocular surface epithelia. METHODS. Full-length mouse Pax6 (mPax) cDNA, or truncated mPax6286 lacking the transcriptional activation domain was inserted into a tetracycline-inducible vector (Tet-on). A rabbit corneal epithelial cell line SIRC was used to establish stable transformants. Induction of Pax6 or truncated Pax6286 proteins by doxycycline (DOX) was examined by Western blot and immunohistochemistry. The effects of Pax6 overexpression on cell cycle progression were assessed by the cell proliferation index, cell growth curve, and cell cycle assay. Terminal deoxynucleotidyl transferase biotin-dUTP nick end labeling (TUNEL) assay was performed to detect apoptotic cells. Recombinant adenovirus-carrying mPax6 or mPax6286 transgenes were used for transient transduction of primary rabbit corneal epithelial cells, and the effect on cell cycle progression was assayed. RESULTS. The level of Pax6 or truncated Pax6 was tightly regulated by DOX. Overexpression of full-length Pax6 retarded the rate of cell proliferation, whereas the truncated form had no effect. Full-length Pax6 affected the rate at which individual cells traversed the cell cycle and induced caspase-3-independent apoptosis in a small percentage of cells. Transient transduction of cells with recombinant mPax6 adenovirus also inhibited cell proliferation. CONCLUSIONS. Inhibition of cell proliferation in Pax6-overexpressing corneal epithelial cell lines and primary cell culture is consistent with the notion that Pax6 plays a role in controlling corneal epithelial cell dynamics in vivo. (Invest Ophthalmol Vis Sci. 2006;47:2397–2407) DOI:10.1167/iovs.05-1083

Pax6 overexpression suppresses cell proliferation and retards the cell cycle in corneal epithelial cells.

Lumican, a small leucine-rich proteoglycan (SLRP), is one of the major extracellular components in interstitial collagenous matrices of the corneal stroma, aorta, skin, skeletal muscle, lung, kidney, bone, cartilage, and intervertebral discs.1–8 In the cornea, lumican contains keratan sulfate chains. However, it is present as a low or nonsulfated glycoprotein (50 –57 kDa) in noncorneal tissues.1,2,9–11 Its wide distribution implies that it has multiple functions in tissue morphogenesis and maintenance of tissue homeostasis. This is best illustrated by the multiple clinical manifestations observed in Lumican-knockout (lum−/−) mice that exhibit corneal opacity, skin and tendon fragility, delayed wound healing, and low fertility. Indeed, lumican has been shown to play essential roles in corneal transparency by regulating collagen fibrillogenesis8 in wound healing, by modulating epithelial cell migration,12 and in the epithelium–mesenchyme transition of the injured lens.13 These results have led to the speculation that lumican may play an active role in corneal epithelial wound healing.

Transplantation of human AM as a temporary or permanent graft has become a popular surgical procedure for ocular surface reconstruction.14,15 Besides the observation that cryopreserved AM reduces inflammation,16–21 scarring,22,23 and neovascularization,24 many clinical studies25–28 have shown that human AM as a graft promotes corneal epithelial wound healing. Furthermore, human AM can help preserve corneal limbal epithelial progenitor cells and maintain the keratocyte phenotype during ex vivo expansion.29–32 Nevertheless, the exact molecular mechanism of the aforementioned effects remains unknown.

The present study revealed that human AM is a rich source of soluble lumican glycoprotein and that purified soluble lumican glycoprotein can facilitate proliferation and migration of corneal epithelial cells during wound healing in both wild-type and lum−/− mice.

/pdf/soluble-lumican-glycoprotein-purified-from-human-amniotic-tmx49f9a0d.pdf

Soluble lumican glycoprotein purified from human amniotic membrane promotes corneal epithelial wound healing.

Keratocan expression of murine keratocytes is maintained on amniotic membrane by down-regulating transforming growth factor-β signaling

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Pax6 overexpression suppresses cell proliferation and retards the cell cycle in corneal epithelial cells.

Soluble lumican glycoprotein purified from human amniotic membrane promotes corneal epithelial wound healing.

Keratocan expression of murine keratocytes is maintained on amniotic membrane by down-regulating transforming growth factor-β signaling