Effects of Thyroid Hormone on Tissue Hypoxia: Relevance to Sepsis Therapy

Tissue hypoxia occurs in various conditions such as myocardial or brain ischemia and infarction, sepsis, and trauma, and induces cellular damage and tissue remodeling with recapitulation of fetal-like reprogramming, which eventually results in organ failure. Analogies seem to exist between the damaged hypoxic and developing organs, indicating that a regulatory network which drives embryonic organ development may control aspects of heart (or tissue) repair. In this context, thyroid hormone (TH), which is a critical regulator of organ maturation, physiologic angiogenesis, and mitochondrial biogenesis during fetal development, may be of important physiological relevance upon stress (hypoxia)-induced fetal reprogramming. TH signaling has been implicated in hypoxic tissue remodeling after myocardial infarction and T3 prevents remodeling of the postinfarcted heart. Similarly, preliminary experimental evidence suggests that T3 can prevent early tissue hypoxia during sepsis with important physiological consequences. Thus, based on common pathways between different paradigms, we propose a possible role of TH in tissue hypoxia after sepsis with the potential to reduce secondary organ failure.

Mucosal Immunology

Approximately 80% of the pathogens that lead to deadly infections in humans choose mucosal tissue as the first site of infection. The mucosal surfaces of the body include the gastrointestinal tract, airways, oral cavity, and urogenital mucosa, which provide a large area conducive to the invasion and accumulation of many microorganisms and are of great importance in this regard. The large extent of mucus, as well as the accumulation of bacteria and countless foreign antigens in these areas, are the most important reasons for the importance of mucosal tissues. In addition to the myriad of symbiotic bacteria, large amounts of oral antigens (both pathogenic and non-pathogenic) enter a person’s body daily and human mucosal tissues are exposed to these antigens. The function of the mucosal immune system is to distinguish pathogenic antigens from non-pathogenic ones. In this way, against a large number of oral antigens or co-tolerant microorganisms, and pathogenic antigens, a favorable (and even non-inflammatory, possible) immune response is produced. Mucosal tissue, as the largest lymphatic organ in the body, is home to 75% of the lymphocyte population and produces the highest amount of immunoglobulin. The amount of secreted IgA (slgA) produced daily by mucosal surfaces is much higher than the IgG produced in the bloodstream. A 70 kg person produces more than 3 grams of IgA per day, which is about 70–60% of the total antibodies produced in the body. The first embryonic organ in which immune system cells are located in the intestine. Some researchers consider this organ (and specifically mucosal lymph nodes) to be the source of the human immune system.

Salivary Gland Organ Regeneration for Oral Health

Salivary glands play essential roles in normal upper gastrointestinal tract function and oral health via saliva production. There are three types of salivary glands; the submandibular gland and parotid gland, which secrete serous saliva, and the sublingual gland secretes mucous saliva in mice [1]. Therefore, salivary gland dysfunction due to atrophy of acinar cells caused by Sjogren’s syndrome, radiation therapy for head and neck cancer, and aging and saliva reduction, and results in xerostomia (dry mouth syndrome), causes various clinical problems in oral health and influences overall bodily health [2]. Xerostomia induces various clinical problems in oral health, such as dental decay, bacterial infection, and disorders of mastication and swallowing, which result in a general reduction in the quality of life [3]. Current standard to cure xerostomia are symptomatic treatments, such as the administration of artificial saliva substitutes and sialogogues, and the administration of parasympathetic stimulation drugs, including pilocarpine and cevimeline [4]. In salivary gland regenerative therapy, stem cell transplantation, which expressed c-kit and Sca-1 as stem cell markers and exhibit high proliferative capacities [5,6], and gene modification for water channel aquaporin-1 (AQP1) and Interleukin-17 (IL-17) receptor antibody is performed to restore the damaged acinar tissue and recover the flow of saliva [7,8]. Salivary glands arise from the organ germ, which is induced by reciprocal epithelial and mesenchymal stem cell interactions during embryonic development [9]. In this two decades, advances in developmental biology have led to break-through in regeneration of functional bioengineered organ in vitro by using embryonic organ-inductive potential stem cells. Recently, regeneration of fully functional salivary gland as a next-generation organ regeneration has been reported after the transplantation of bioengineered salivary gland germ developed by using embryonic organ-inductive potential stem cells as well as iPS cells [10,11]. In 2013, we demonstrated the fully functional salivary gland regeneration by recapitulating the embryonic morphogenesis. The bioengineered salivary gland organ germ, which was reconstituted by our Organ Germ Method using organ-inductive potential stem cells derived from submandibular gland germs of ED 13.5 mice [12], showed reciprocal epithelial and mesenchymal interactions in in vitro organ culture. Following the orthotopic transplantation, the bioengineered germ develops into a mature salivary gland via acinar formations with a myoepithelium and innervation. The bioengineered salivary gland produces saliva in response to the pilocarpine administration and gustatory stimulation by citrate, protected against bacterial infection and restores normal swallowing function in a mouse model. This study provides a proofof- concept for bioengineered salivary gland regeneration. The next breakthrough first reported in 2008 was the emergence of an organoid as a mini-organ that was generated by inducing body patterning and a subsequent organ-forming field formation in pluripotent stem cells [13]. Organoids are useful to recapitulate the process of organogenesis, which are strictly regulated by morphogen signalling and transcriptional networks. To date, multiple types of organoids, including the retina, pituitary gland, liver, and lung have been successfully generated in vitro. Our colleagues identified a specific combination of two transcription factors, Sox9 and Foxc1, are responsible for the differentiation of mouse ES cells-derived oral ectodermal region into the salivary gland rudiment in a threedimensional organoid culture system [11]. The induced salivary gland rudiment showed a similar morphologies and gene expression profiles to those of the embryonic salivary gland rudiment of normal mice. Following orthotopic transplantation into salivary glands-defected mice, the induced salivary gland germ exhibited characteristics of mature salivary glands including histological morphologies, nerve innervation and saliva secretion. This study is the first report of the fully functional organ regeneration by using organoid, demonstrating the potential of salivary gland organ regeneration from pluripotent stem cells for an additional organ replacement regenerative therapy. As part of regenerative medicine, the autogenous transplantation of stem cells, including bone marrow, and tissue sheets such as skin, cornea, and cardiac muscle, has already been applied clinically through cell and tissue transplantation therapies. The progress made in the past two decades has been remarkable, paving the way for possible future organ replacement regenerative therapies. There remains a critical issue as to whether next-generation organ regenerative therapy can be adopted as a novel clinical therapy for patients of the loss of organ function. The bioengineered salivary gland organ germ by using both embryonic organ-inductive potential stem cells and pluripotent stem cells can grow where salivary grand should be in oral cavity and produce the saliva in response to various taste stimulations. These works indicate the potentials for clinical application of salivary gland organ regeneration. Applying knowledge of these basic research will enable the regeneration of salivary gland organ in the next decades.

Exploiting teeth as a model to study basic features of signaling pathways

Teeth constitute a classical model for the study of signaling pathways and their roles in mediating interactions between cells and tissues in organ development, homeostasis and regeneration. Rodent teeth are mostly used as experimental models. Rodent molars have proved fundamental in the study of epithelial–mesenchymal interactions and embryonic organ morphogenesis, as well as to faithfully model human diseases affecting dental tissues. The continuously growing rodent incisor is an excellent tool for the investigation of the mechanisms regulating stem cells dynamics in homeostasis and regeneration. In this review, we discuss the use of teeth as a model to investigate signaling pathways, providing an overview of the many unique experimental approaches offered by this organ. We discuss how complex networks of signaling pathways modulate the various aspects of tooth biology, and the models used to obtain this knowledge. Finally, we introduce new experimental approaches that allow the study of more complex interactions, such as the crosstalk between dental tissues, innervation and vascularization.

Identifying stem cell numbers and functional heterogeneities during post-embryonic organ growth

SummaryUncovering the number of stem cells necessary to grow an organ has been challenging in most vertebrate systems. Here, we have developed a mathematical model that we use to characterise stem cells in the fish gill, an organ that displays non-exhaustive growth. Our work employs a Markov model, first stochastically simulated via an adapted Gillespie algorithm, and further improved by using probability theory. The stochastic algorithm produces a simulated data set for comparison with experimental data by inspecting quantifiable properties, while the analytical approach skips the step of in silico data generation and goes directly to the quantification, being more abstract and very efficient. By applying the model to a large clonal experimental dataset, we report that a reduced number of stem cells are responsible for growing and maintaining the fish gill. The model also highlights a functional heterogeneity among the stem cells involved, where activation and quiescence phases determine their relative growth contribution. Overall, our work presents an easy-to-apply algorithm to infer the number of stem cells functionally required in a life-long growing system.

Quenching autofluorescence in tissue immunofluorescence

Background: Immunofluorescence (IF) is one of the most important techniques where fluorochromes conjugated to antibodies are used to detect specific proteins or antigens. In tissue sections, autofluorescence (AF) can lead to poor quality images that impair assessment. The placenta is a pivotal extra-embryonic organ in embryo development, where trophoblasts make up a large proportion of the cells. Teratoma formation is one of the critical assays for pluripotent stem cells. Methods: We tested whether ultraviolet (UV), ammonia (NH3), copper (II) sulfate (CuSO4), Trypan Blue (TB), Sudan Black B (SB), TrueBlack™ Lipofusin Autofluorescence Quencher (TLAQ) and combinations of these treatments could reduce AF in paraffin and frozen sections of placenta and teratoma in FITC, Texas Red and Cy5.5 channels. Results: We found that UV, NH3, TB and CuSO4 quenched AF to some extent in different tissue and filters, but increased AF in Texas Red or Cy5.5 channels in some cases. SB and TLQA exhibited the most consistent effects on decreasing AF, though TLQA reduced the overall IF signal in placenta sections. Not all combined treatments further reduced AF in both placenta and teratoma sections. Conclusions: SB and TLAQ can effectively quench AF in placenta and teratoma IF.