Construction and composition of the squid pen from Doryteuthis pealeii

Author Posting. © University of Chicago Press, 2019. This article is posted here by permission of University of Chicago Press for personal use, not for redistribution. The definitive version was published in Messerli, M. A., Raihan, M. J., Kobylkevich, B. M., Benson, A. C., Bruening, K. S., Shribak, M., Rosenthal, J. J. C., & Sohn, J. J. Construction and composition of the squid pen from Doryteuthis pealeii. Biological Bulletin. 237(1), (2019): 1-15, doi:10.1086/704209.The pen, or gladius, of the squid is an internalized shell. It serves as a site of attachment for important muscle groups and as a protective barrier for the visceral organs. The pen’s durability and flexibility are derived from its unique composition of chitin and protein. We report the characterization of the structure, development, and composition of pens from Doryteuthis pealeii. The nanofibrils of the polysaccharide β-chitin are arranged in an aligned configuration in only specific regions of the pen. Chitin is secreted early in development, enabling us to characterize the changes in pen morphology prior to hatching. The chitin and proteins are assembled in the shell sac surrounded by fluid that has a significantly different ionic composition from squid plasma. Two groups of proteins are associated with the pen: those on its surface and those embedded within the pen. Only 20 proteins are identified as embedded within the pen. Embedded proteins are classified into six groups, including chitin associated, protease, protease inhibitors, intracellular, extracellular matrix, and those that are unknown. The pen proteins share many conserved domains with proteins from other chitinous structures. We conclude that the pen is one of the least complex, load-bearing, chitin-rich structures currently known and is amenable to further studies to elucidate natural construction mechanisms using chitin and protein.We thank John Dowling for financial support. We thank Kasia Hammar and Louie Kerr of the Marine Biological Laboratory Central Microscopy Facility for help obtaining scanning electron micrographs. We thank Bogdan Budnik and Renee Robinson from the Mass Spectrometry and Proteomics Resource Laboratory for their help and advice with protein identification. We thank Shin-Yi Marzano and Chenchen Feng of South Dakota State University for help with rapid amplification of cDNA ends. Funding for this work was provided by the Eugene and Millicent Bell Fellowship Fund in Tissue Engineering (MAM), an Agriculture and Biological Sciences Undergraduate Research Award (KSB), National Institutes of Health grant R01 GM101701 (MS), National Science Foundation grant IOS1557748 (JJCR), and Israel-United States Binational Science Foundation 2013094 (JJCR). Literature Cited2020-07-0

Bovine Cellular and Innate Immune Response to Hemostatic Polysaccharides

Chronic wounds are longstanding dermal wounds that do not heal in a normal amount of time and can commonly last for over a year. These wounds plague humans and farm animals alike. Wounds are characterized by chronic inflammation and are often painful, difficult to heal, have a high recurrence rate, and are potentially debilitating. Long term wounds that do not fully heal are distressing for both humans and animals, but understandably more resources have been directed towards humans. Some treatment options for humans may be suitable for adaptation into the field of animal care. Chronic lesions such as bovine digital dermatitis (BDD), sole ulcers, and foot rot might be treated using proven techniques from human medicine. BDD is a chronic wound, characterized by poly-microbial infection of the hoof that is highly transmissible between cattle. BDD has been reported to greatly impact milk production in dairy herds, result in lameness, and require potentially costly preventative and/or treatment methods. Polysaccharide bandages, currently used to stop hemorrhaging in humans, serve as a potential treatment for chronic wounds. Oxidized regenerated cellulose (ORC) and chitin or chitosan, are effective hemostatic bandages that are being explored to promote wound healing. These novel materials promote some beneficial effects in human wounds but our knowledge of their effects on other mammals is nearly non-existent. We have examined the innate immune response of bovine dermal cells to these polysaccharide hemostats. Polysaccharide materials were characterized with cross polarization-nuclear magnetic resonance (CP-NMR) prior to experimentation. The innate immune response of bovine fibroblasts was monitored by measuring mRNA expression of select cytokines and Toll-like receptors after exposure to ORC and chitin for 24 hours. The roles of ORC and chitin on the lipopolysaccharide (LPS) induced inflammatory response, and fibroblast viability were also determined The results demonstrated that both ORC and chitin elicit a relatively weak immune response from the fibroblasts, reduce expression of inflammatory cytokines in response to LPS, and do not increase mortality at the concentrations used. In conjunction with other previously characterized properties these polysaccharides may improve the healing of chronic epidermal wounds in humans and farm animals