Hyaluronan in cell protrusions and microvesicles, studies on their structure, function and existence in vivo

Public examination of a doctoral dissertation in the field of Cell Biology

Doctoral candidate: Lic Med Ville Koistinen

Date and venue: 21.6.2017 at 12 noon, Snellmania, SN200, Kuopio Campus

Language of the public examination: Finnish

Language of the dissertation: English

Hyaluronan is a large unbranched glycosaminoglycan. The molecular mass of a single molecule can reach 10 million daltons and a stretched length of up to 25 mm. Hyaluronan is very hydrophilic due to a negative charge and it forms a highly viscous solution in water.  While hyaluronan has an important role as a space filler in tissues, it also contributes to essential cellular processes, including embryonic development, cell migration, proliferation, wound healing and cancer progression.

Hyaluronan is synthesized by three plasma membrane proteins, called hyaluronan synthases (HAS1-3). Previous studies on cultured cells have shown that overexpression of HAS2 and HAS3 induce the formation of extensive microvilli. The microvilli are covered with a hyaluronan coat and are dependent on filamentous actin. Nevertheless, several questions about the microvilli remain; do these structures exist in vivo? What is the exact role of hyaluronan in their maintenance? What might be their function? Therefore, the aim of this dissertation was to study the ultrastructure and function of the hyaluronan coated microvilli in more detail, and to investigate if these special structures exist in vivo.

The results show that HAS overexpression drives the assembly of actin filaments in the cell cortex and into the protrusions. The core of the HAS-induced microvilli consist of only  ~8 actin microfilaments which implies that in order to maintain such long and slender protrusions, additional support is needed. Indeed, the hyaluronan coat around the microvilli acts as an extracellular cytoskeleton. Therefore, active hyaluronan synthesis is required for the growth and maintenance of the microvilli. The activity of hyaluronan synthesis was also found to correlate with the shedding of microvesicles, covered by hyaluronan, and budded off from the tips of the microvilli.

The HAS-induced microvilli in cultured cells, and the microvilli on mesothelial surfaces were ultrastructurally similar. Mesothelium was also positive for hyaluronan, but negative for the main hyaluronan receptor CD44. However, epithelial to mesenchymal transition induced by mesothelial wounding or epidermal growth factor increased hyaluronan synthesis, the expression of CD44 and HASs, the formation of microvilli and the shedding of microvesicles. These findings suggest that the HAS-induced microvilli serve as a source of microvesicles, a recently discovered vehicle for the transport of signals in the regulation of wound healing and progression of cancer. CD44 positive microvesicles could serve as a source for new biomarkers to detect EMT-related processes in tissue injuries and cancer.

In the future, the HAS-induced microvilli and microvesicles may prove to be a useful tool in clinical applications such as cancer diagnosis and therapy.

The doctoral dissertation of Ville Koistinen, Licentiate of Medicine, entitled Hyaluronan in cell protrusions and microvesicles, studies on their structure, function and existence in vivo will be examined at the Faculty of Health Sciences. The Opponent in the public examination will be Professor Pekka Lappalainen of the University of Helsinki, and the Custos will be Docent Kirsi Rilla of the University of Eastern Finland.

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