Human pluripotent stem cells : glycomic approaches for culturing and characterization

Human pluripotent stem cells (including embryonic stem cells and induced pluripotent stem cells) are defined by two important characteristics: unlimited self-renewal capacity and ability to switch on various differentiation pathways. These unique properties make them valuable tools for basic research and for the development of regenerative therapies. Since the discovery of stem cells, diverse culture conditions have been developed. Pluripotent stem cells are cultured either in the presence of feeder cells or with extracellular matrix components that together with growing colonies create a niche supporting the growth of undifferentiated cells. Current standard in vitro cell culture techniques are based on the use of xenogeneic reagents. However, this is not compatible with the clinical applications of human pluripotent stem cells. The aim of this study was to develop optimal culture conditions for pluripotent stem cells without the use of xenogeneic reagents, based on the analysis of their cell surface glycan expression. Initially, postnatal human feeder cells, foreskin fibroblasts, were shown to support the derivation of new embryonic stem cell lines and continued undifferentiated growth of these cells. However, the growth rate of stem cells was significantly lower on human feeder cells than on mouse embryonic fibroblasts. This feature restricts the use of human feeder cells for large-scale cell production of stem cells. Next, a number of human embryonic stem cell lines were derived and characterized in detail. The results show that despite of similar basic characteristics in the undifferentiated state, the differentiation capacity of stem cell lines varies. This highlights the need to identify markers that reliably predict cell lineage propensity of the stem cells lines. To identify such predictive markers we conducted a global analysis of cell surface glycans expressed on pluripotent human stem cells. The results show that embryonic stem cells have a unique glycan fingerprint that differs from their differentiated derivatives. This suggests that information of stem cell surface glycans can be used to design markers that define specific stages of differentiation. In addition, this information can be used to develop defined reagents for stem cell culture. Based on the analysis of stem cell surface glycans, specific glycan-binding lectins were studied for their capacity to support human pluripotent stem cells. This lead to the discovery of a lectin, Erythrina Cristagalli (ECA) as a potent simple defined matrix for human pluripotent stem cells. This simple defined matrix, combined with a defined culture medium and the use of a Rho-kinase inhibitor at the time of cell propagation, allowed more efficient production of high-quality human pluripotent stem cells than Matrigel®, the current standard acellular matrix used for stem cell culture. Taken together, these studies advance the development of technologies needed for the efficient generation of fully undifferentiated human pluripotent stem cells in defined conditions.Pluripotenteilla kantasoluilla on kaksi erityispiirrettä: lähes rajaton jakautumiskyky ja kyky erilaistua moniksi erilaistuneiksi solutyypeiksi. Näiden ominaisuuksien ansiosta kantasoluilla on useita mahdollisia lääketieteellisiä ja farmakologisia sovelluskohteita. Tässä työssä on tutkittu ihmisen pluripotenttien kantasoluviljelmien erityisominaisuuksia ja viljelmien välisiä eroavaisuuksia niiden kyvyssä kasvaa ja erilaistua. Lisäksi on tutkittu kantasolujen solupinnan sokerirakenteita ja kehitetty sokerirakenteisiin perustuvia viljelymenetelmiä. Ensimmäisen osajulkaisun tavoitteena oli kehittää ihmisperäisten pluripotenttien solujen viljelyolosuhteita, joissa vältetään toisesta lajista (hiirestä) peräisin olevia komponentteja. Työssä osoitettiin ihmisperäisten fibroblastisolujen kykenevän toimimaan ihmisen alkion kantasolujen tukisoluina. Toisessa osatyössä kuvattiin viiden alkion kantasolulinjan tuotto ja karakterisaatio. Työssä tutkittiin myös viljelyolosuhteiden vaikutusta solujen kasvuun, kantasolulinjojen ilmentämiä geenituotteita ja erilaistumiskykyä. Tutkimuksessa havaittiin alkion kantasolujen muistuttavan geeni-ilmenemiseltään sukusolujen kantasoluja. Lisäksi osoitettiin kantasolulinjojen poikkeavan toisistaan erilaistumiskyvyltään huolimatta niiden samankaltaisesta geeni-ilmenemisestä. Kolmannessa osatyössä tutkittiin kantasolujen ilmentämiä N-glykaani sokerirakenteita. Työssä havaittiin massaspektrometrian avulla pluripotenteilla kantasoluilla oleva joukko niille tyypillisiä sokerirakenteita, jotka muuttuvat kantasolujen erilaistuessa. Neljännessä osatyössä kehitettiin kantasolujen viljelymenetelmiä hyödyntäen kolmannessa osatyössä havaittuja kantasolujen solupinnan sokerirakenteita ja osoitetaan erään lektiinin (Erythrina cristagalli) tukevan kantasolujen kiinnittymistä, kasvua ja kantasoluominaisuuksien säilymistä. Tämä työ osoittaa, että viljelyolosuhteiden kehittäminen ja validoiminen on ensiarvoisen tärkeää pluripotenttien kantasolulinjojen tutkimukselle ja lääke-tieteelliselle käytölle

Fearfulness associates with problematic behaviors and poor socialization in cats

Funding Information: This study was funded by Suomen Kissaliitto ry , Kinship Partners Inc , Academy of Finland ( 308887 ), and Agria and Swedish Kennel Club research fund ( N2019-0005 ). We acknowledge Julia Niskanen for developing the airGLMs for a model selection process. Publisher Copyright: © 2022 The Author(s)Problematic behavior is a remarkable welfare issue in cats (Felis catus), as it is one of the most common reasons for relinquishment. The probability of developing problematic behaviors is likely influenced by several variables, but these remain little studied. In this study, we examined the associations of fearfulness, aggression toward humans, and excessive grooming with nearly thirty variables in a survey dataset of over 3,200 cats. To identify the most important variables influencing these behaviors, we used generalized linear models. All behaviors were associated with each other suggesting comorbidity between problematic behaviors. Breed and several environmental variables were also associated with behaviors. Poor socialization with humans and a history of being a rescue cat were associated with higher fearfulness, indicating that the proper socialization of kittens is beneficial for avoiding fear-related problematic behaviors. Overall, our study highlights the complexity of three problematic behaviors in cats.Peer reviewe

Habitat selection of the mud crab Rhithropanopeus harrisii in its newly invaded range

Information on the habitat selection by non-indigenous species is crucial for understanding their effects on the communities to which they are introduced, since the effects are often focused on the invaded habitats. The North American mud crab Rhithropanopeus harrisii is a new invader in the northern Baltic Sea, on the coasts of Finland and Estonia. In the Finnish Archipelago Sea, it has been found in two very distinct habitats: reed belts of Phragmites australis and algal zones with Fucus vesiculosus as the main habitat-forming species. In previous studies in the Baltic Sea, R. harrisii has preferred F. vesiculosus and has locally driven a shift in the structure of F. vesiculosus-associated invertebrate communities. Here, we disentangled whether habitat choice was determined by habitat structure or the availability of food. First, we conducted a habitat selection experiment with P. australis and F. vesiculosus habitats and varying food availability, and found that R. harrisii preferred F. vesiculosus, with food having no effect on the habitat choice. Second, we studied if the preference for F. vesiculosus was due to the alga itself or the rocks it grows on. We found that R. harrisii preferred the shelter of the rock habitat, indicating that R. harrisii choose their habitat based on habitat structure rather than food availability in the habitat. However, the preference for sheltered rocky bottom habitats also exposes the associated F. vesiculosus communities to the impacts of R. harrisii through predation.Peer reviewe

Personality traits associate with behavioral problems in pet dogs

Personality traits, especially neuroticism, strongly predict psychopathology. The domestic dog (Canis lupus familiaris Linnaeus, 1758) is used as a natural model for psychiatric disorders, but the similarity between dog and human personality and the association between dog personality and unwanted behavioral traits, such as fearfulness, aggressiveness, and impulsivity/inattention, remain unknown. This study utilized structural equation modeling (SEM) with survey data of 11,360 dogs to examine the associations and correlations between seven personality and ten unwanted behavioral traits. Personality traits included insecurity, energy, training focus, aggressiveness/dominance, human sociability, dog sociability, and perseverance. Unwanted behavioral traits included fearfulness, noise sensitivity, fear of surfaces/heights, separation anxiety, barking, stranger-directed aggression, owner-directed aggression, dog-directed aggression, hyperactivity/impulsivity, and inattention. We first fitted confirmatory factor models for the unwanted behavioral traits and the best model grouped unwanted behaviors into four latent traits: fear-related behavior, fear-aggression, aggression, and impulsivity/inattention and used this structure in the subsequent SEM model. Especially, insecurity, which resembles the human neuroticism trait, was strongly associated with unwanted behavior, paralleling the association between neuroticism and psychopathology. Similarly, training focus, resembling conscientiousness, was negatively related to impulsivity/inattention, and aggressiveness/dominance was associated with aggressive behaviors, resembling associations of conscientiousness and agreeableness with attention deficit hyperactivity disorder and aggression-related psychopathology, respectively. These results indicate that dog personality traits resemble human personality traits, suggesting that their neurological and genetic basis may also be similar and making the dog a suitable animal model for human behavior and psychiatric disorders.Peer reviewe

Aggressive behaviour is affected by demographic, environmental and behavioural factors in purebred dogs

Aggressive behaviour is an unwanted and serious problem in pet dogs, negatively influencing canine welfare, management and public acceptance. We aimed to identify demographic and environmental factors associated with aggressive behaviour toward people in Finnish purebred pet dogs. We collected behavioural data from 13,715 dogs with an owner-completed online questionnaire. Here we used a dataset of 9270 dogs which included 1791 dogs with frequent aggressive behaviour toward people and 7479 dogs without aggressive behaviour toward people. We studied the effect of several explanatory variables on aggressive behaviour with multiple logistic regression. Several factors increased the probability of aggressive behaviour toward people: older age, being male, fearfulness, small body size, lack of conspecific company, and being the owner's first dog. The probability of aggressive behaviour also differed between breeds. These results replicate previous studies and suggest that improvements in the owner education and breeding practices could alleviate aggressive behaviour toward people while genetic studies could reveal associated hereditary factors.Peer reviewe

Aggressiveness, ADHD-like behaviour, and environment influence repetitive behaviour in dogs

Repetitive behaviour ranges from variants of normal repetitive behaviours to abnormal repetitive behaviours. The domestic dog spontaneously performs different repetitive behaviours, which can be severe and impair the quality of life and the dog-owner relationship. We collected comprehensive behavioural questionnaire data from almost 4500 Finnish pet dogs and studied the effect of several demographic, environmental, and behavioural factors on canine repetitive behaviour with logistic regression. We replicated findings from previous studies by revealing comorbidity between repetitive behaviour and behavioural factors aggressiveness, hyperactivity/impulsivity, and inattention. We also found a novel association between repetitive behaviour and the owner's dog experience. In addition, we showed that repetitive behaviour is more common in dogs that live without conspecifics, dogs that were given a low amount of exercise, dogs that lived in larger families, young dogs and elderly dogs, and neutered dogs. Finally, we identified breed differences in repetitive behaviour, suggesting that some breeds are more vulnerable to repetitive behaviour and indicate a genetic susceptibility. As abnormal repetitive behaviour can considerably worsen the well-being of dogs and impair the dog-owner relationship, a better understanding of the environmental, lifestyle, and molecular factors affecting canine repetitive behaviour can benefit both dogs and humans.Peer reviewe

Active and social life is associated with lower non-social fearfulness in pet dogs

Behavioural problems are leading welfare issues in domestic dogs. In particular, anxiety-related behavioural problems, such as fearfulness and noise sensitivity are highly prevalent conditions that cause distress to fearful dogs. To better understand the environmental factors associated with non-social fear, including noise sensitivity, fear of novel situations, and fear of surfaces and heights, a large online survey including data on 13,700 Finnish pet dogs was performed by the dog owners. After fulfilling inclusion criteria, this data consisted of 9,613 dogs with fear of fireworks, 9,513 dogs with fear of thunder, 6,945 dogs with fear of novel situations, and 2,932 dogs with fear of surfaces and heights. Logistic regression analyses revealed that dogs with frequent non-social fear had experienced less socialisation during puppyhood, were more often neutered, had inexperienced owners, lived without conspecifics, participated less frequently in activities or training, and lived in more urban environments. In addition, we identified several breed differences, and a tendency of more common non-social fear in small dog breeds, which suggests a genetic background. Non-social fearfulness has a negative effect on well-being of the dogs. Our findings suggest that the socialisation and the living environment and the value of other dogs' company and owner interaction via activities and training may improve the well-being of the dogs.Peer reviewe

Inadequate socialisation, inactivity, and urban living environment are associated with social fearfulness in pet dogs

Problematic behaviours are severe welfare issues for one of the world's most popular pets, the domestic dog. One of the most prevalent behavioural problem that causes distress to dogs is social fearfulness, meaning fear of conspecifics or unfamiliar people. To identify demographic and environmental factors associated with fear of dogs and strangers, logistic regression was utilised with a large dataset of 6,000 pet dogs collected through an owner-filled behavioural survey. Social fearfulness was associated with several factors, including urban environment, poor socialisation during puppyhood, infrequent participation in training and other activities, small body size, female sex, and neutering. In addition, we identified several breed differences, suggesting a genetic contribution to social fearfulness. These findings highlight the role of inadequate socialisation, inactivity, and urban living environmental in fear-related behavioural problems in dogs. Improvements in the management and breeding practices of dogs could, therefore, enhance the welfare of man's best friend.Peer reviewe

Habitat selection of the mud crab Rhithropanopeus harrisii in its newly invaded range

Information on the habitat selection by non-indigenous species is crucial for understanding their effects on the communities to which they are introduced, since the effects are often focused on the invaded habitats. The North American mud crab Rhithropanopeus harrisii is a new invader in the northern Baltic Sea, on the coasts of Finland and Estonia. In the Finnish Archipelago Sea, it has been found in two very distinct habitats: reed belts of Phragmites australis and algal zones with Fucus vesiculosus as the main habitat-forming species. In previous studies in the Baltic Sea, R. harrisii has preferred F. vesiculosus and has locally driven a shift in the structure of F. vesiculosus-associated invertebrate communities. Here, we disentangled whether habitat choice was determined by habitat structure or the availability of food. First, we conducted a habitat selection experiment with P. australis and F. vesiculosus habitats and varying food availability, and found that R. harrisii preferred F. vesiculosus, with food having no effect on the habitat choice. Second, we studied if the preference for F. vesiculosus was due to the alga itself or the rocks it grows on. We found thatR. harrisii preferred the shelter of the rock habitat, indicating that R. harrisii choose their habitat based on habitat structure rather than food availability in the habitat. However, the preference for sheltered rocky bottom habitats also exposes the associated F. vesiculosus communities to the impacts of R. harrisii through predation. </p