Biology and ecology of tardigrades (Ecdysozoa: Tardigrada)

Ovaj završni seminarski rad bavi se općenitim prikazom biologije i ekologije dugoživaca. Dugoživci (Tardigrada) su protostomični beskralježnjaci mikroskopske veličine od 0,1 do 2 mm što ih svrstava u meiofaunu. Iako su im pripisivane različite sistematske kategorije, danas se smatraju pripadnicima nadkoljena Ecdysozoa. Opisano je preko tisuću vrsta podijeljenih u tri razreda. Relativno su jednostavne tjelesne građe slične onoj u člankonožaca, kolutićavaca i crvonožaca. Najpoznatiji su po sposobnosti ulaska u stanje kriptobioze, tj. preživljavanja niskih temperatura, suše, visokih doza zračenja i sl., iako to mogu i neke druge vrste. Evolucija dugoživaca nije u potpunosti jasna ali je otkriveno da su srodstveno ipak bliži crvonošcima nego oblićima. Geografski su široko rasprostranjeni po kopnenim, morskim i slatkovodnim biotopima. Njihova najpoznatija i najbolje istražena staništa su mahovine i lišajevi, no nađeni su i na ekstremnim staništima poput ledenjaka ili termalnih izvora. Ekološka uloga, kao i drugim pripadnicima meiofaune, im nije beznačajna što je dokaz kompleksnosti trofičkih odnosa na raznim mikrostaništima. Tardigradološka istraživanja u zadnjih deset godina su pojačana što je dokaz da dugoživci nisu opskurno, već manje poznato koljeno živog svijeta te da su izazov ne samo za klasičnu zoologiju već za fundamentalna istraživanja u znanosti o životu.This undergraduate thesis finds its purpose in presenting general review of tardigrade biology and ecology. Tardigrades (Tardigrada) are protostomic invertebrates of microscopic size varying from 0,1 to 2 mm, what places them in meiofauna. Formerly they were assigned various systematic positions, but today tardigrades are considered members of Ecdysozoa superphylum. There are over thousand described species divided into three classes. Their fairly simple anatomy is similar to that found in arthropods, annelids and onychophorans. Cryptobiosis, i. e. surviving low temperatures, drought and high doses of radiation is the best known remark of tardigrades. Evolution of Tardigrada still remains partialy unclear, but phylogenetic analyses have proven their closer relationship to onychophorans than nematodes. Tardigrades have cosmopolitan distribution which extends from marine and freshwater, to terrestrial biotopes. Although mosses and lichens are the most common and best investigated tardigrade habitats, they were also recovered from glacier ice and hot springs. Ecological role of tardigrades, as well as other meiofauna members is is not negligible, and, in fact, proves complex trophic relationships on a broad range of microhabitats. Tardigradological investigations have been intensified during the last decade. Once again, that is a reminder tardigrades are not obscure, but less-known phylum of the living world, posing a challenge not only for classical zoology, but for fundamental life science research

Biology and ecology of tardigrades (Ecdysozoa: Tardigrada)

Ovaj završni seminarski rad bavi se općenitim prikazom biologije i ekologije dugoživaca. Dugoživci (Tardigrada) su protostomični beskralježnjaci mikroskopske veličine od 0,1 do 2 mm što ih svrstava u meiofaunu. Iako su im pripisivane različite sistematske kategorije, danas se smatraju pripadnicima nadkoljena Ecdysozoa. Opisano je preko tisuću vrsta podijeljenih u tri razreda. Relativno su jednostavne tjelesne građe slične onoj u člankonožaca, kolutićavaca i crvonožaca. Najpoznatiji su po sposobnosti ulaska u stanje kriptobioze, tj. preživljavanja niskih temperatura, suše, visokih doza zračenja i sl., iako to mogu i neke druge vrste. Evolucija dugoživaca nije u potpunosti jasna ali je otkriveno da su srodstveno ipak bliži crvonošcima nego oblićima. Geografski su široko rasprostranjeni po kopnenim, morskim i slatkovodnim biotopima. Njihova najpoznatija i najbolje istražena staništa su mahovine i lišajevi, no nađeni su i na ekstremnim staništima poput ledenjaka ili termalnih izvora. Ekološka uloga, kao i drugim pripadnicima meiofaune, im nije beznačajna što je dokaz kompleksnosti trofičkih odnosa na raznim mikrostaništima. Tardigradološka istraživanja u zadnjih deset godina su pojačana što je dokaz da dugoživci nisu opskurno, već manje poznato koljeno živog svijeta te da su izazov ne samo za klasičnu zoologiju već za fundamentalna istraživanja u znanosti o životu.This undergraduate thesis finds its purpose in presenting general review of tardigrade biology and ecology. Tardigrades (Tardigrada) are protostomic invertebrates of microscopic size varying from 0,1 to 2 mm, what places them in meiofauna. Formerly they were assigned various systematic positions, but today tardigrades are considered members of Ecdysozoa superphylum. There are over thousand described species divided into three classes. Their fairly simple anatomy is similar to that found in arthropods, annelids and onychophorans. Cryptobiosis, i. e. surviving low temperatures, drought and high doses of radiation is the best known remark of tardigrades. Evolution of Tardigrada still remains partialy unclear, but phylogenetic analyses have proven their closer relationship to onychophorans than nematodes. Tardigrades have cosmopolitan distribution which extends from marine and freshwater, to terrestrial biotopes. Although mosses and lichens are the most common and best investigated tardigrade habitats, they were also recovered from glacier ice and hot springs. Ecological role of tardigrades, as well as other meiofauna members is is not negligible, and, in fact, proves complex trophic relationships on a broad range of microhabitats. Tardigradological investigations have been intensified during the last decade. Once again, that is a reminder tardigrades are not obscure, but less-known phylum of the living world, posing a challenge not only for classical zoology, but for fundamental life science research

Biology and ecology of tardigrades (Ecdysozoa: Tardigrada)

Ovaj završni seminarski rad bavi se općenitim prikazom biologije i ekologije dugoživaca. Dugoživci (Tardigrada) su protostomični beskralježnjaci mikroskopske veličine od 0,1 do 2 mm što ih svrstava u meiofaunu. Iako su im pripisivane različite sistematske kategorije, danas se smatraju pripadnicima nadkoljena Ecdysozoa. Opisano je preko tisuću vrsta podijeljenih u tri razreda. Relativno su jednostavne tjelesne građe slične onoj u člankonožaca, kolutićavaca i crvonožaca. Najpoznatiji su po sposobnosti ulaska u stanje kriptobioze, tj. preživljavanja niskih temperatura, suše, visokih doza zračenja i sl., iako to mogu i neke druge vrste. Evolucija dugoživaca nije u potpunosti jasna ali je otkriveno da su srodstveno ipak bliži crvonošcima nego oblićima. Geografski su široko rasprostranjeni po kopnenim, morskim i slatkovodnim biotopima. Njihova najpoznatija i najbolje istražena staništa su mahovine i lišajevi, no nađeni su i na ekstremnim staništima poput ledenjaka ili termalnih izvora. Ekološka uloga, kao i drugim pripadnicima meiofaune, im nije beznačajna što je dokaz kompleksnosti trofičkih odnosa na raznim mikrostaništima. Tardigradološka istraživanja u zadnjih deset godina su pojačana što je dokaz da dugoživci nisu opskurno, već manje poznato koljeno živog svijeta te da su izazov ne samo za klasičnu zoologiju već za fundamentalna istraživanja u znanosti o životu.This undergraduate thesis finds its purpose in presenting general review of tardigrade biology and ecology. Tardigrades (Tardigrada) are protostomic invertebrates of microscopic size varying from 0,1 to 2 mm, what places them in meiofauna. Formerly they were assigned various systematic positions, but today tardigrades are considered members of Ecdysozoa superphylum. There are over thousand described species divided into three classes. Their fairly simple anatomy is similar to that found in arthropods, annelids and onychophorans. Cryptobiosis, i. e. surviving low temperatures, drought and high doses of radiation is the best known remark of tardigrades. Evolution of Tardigrada still remains partialy unclear, but phylogenetic analyses have proven their closer relationship to onychophorans than nematodes. Tardigrades have cosmopolitan distribution which extends from marine and freshwater, to terrestrial biotopes. Although mosses and lichens are the most common and best investigated tardigrade habitats, they were also recovered from glacier ice and hot springs. Ecological role of tardigrades, as well as other meiofauna members is is not negligible, and, in fact, proves complex trophic relationships on a broad range of microhabitats. Tardigradological investigations have been intensified during the last decade. Once again, that is a reminder tardigrades are not obscure, but less-known phylum of the living world, posing a challenge not only for classical zoology, but for fundamental life science research

Changes in subclass-specific IgG Fc glycosylation associated with the postnatal maturation of the murine immune system

Early postnatal life is characterized by a critical time period in which the developing neonatal immune system transitions from passive immunity, induced by protective maternal antibodies, to the competence of a fully functioning immune system. The inflammatory capability of both maternal and neonatal antibodies is governed by N-linked glycosylation of the Fc region, and though this has been examined extensively in adults, there is currently little information regarding antibody glycosylation patterns during early postnatal life. To characterize the murine IgG Fc glycosylation profile during early life, we used nano-LC-ESI-Qq-TOF mass spectrometry analysis to assess subclass specific Asn-297 glycosylation patterns in the serum of BALB/c mice from 5–60 days of age. From birth to adulthood, we observed a decline in proinflammatory Fc glycosylation in all IgG subclasses. This was shown by significantly reduced agalactosylated and monogalactosylated structures combined with increased sialylation after weaning at 45 and 60 days of age. This information indicates that the transition between neonatal life and adulthood in mice is accompanied by reduction of inflammatory IgG antibodies. Our study contributes to a growing body of literature indicating the importance of IgG Fc glycosylation and its association with inflammation during different life stages

Properties and function of mucous coat on the cuticle of armoured tardigrades (Tardigrada: Heterotardigrada)

Dugoživci su mikroskopski beskralježnjaci koji nepovoljne uvjete u okolišu, poput isušivanja, preživljavaju ulaskom u reverzibilno ametaboličko stanje kriptobioze. Tijelo im prekriva višeslojna kutikula na čijoj je površini mukozni sloj. Cilj ovog rada bio je odrediti sastav, raspored, morfologiju i nanomehanička svojstva mukoznog sloja na kutikuli oklopljenog dugoživca Echiniscus testudo tehnikama bojenja Alcian-modrilom, koloidnim željezom, Nilskim crvenilom i Coomassie briljant plavim, Ramanovom spektroskopijom, pretražnom elektronskom mikroskopijom i mikroskopijom atomskih sila. Materijal mukoznog sloja, u kojem su dokazani mukopolisaharidi, lipidi i proteini, lokaliziran je pretežno u udubljenjima i zonama pregiba dorzalne kutikule. Mikroskopijom atomskih sila oslikana je površina suhe i hidratizirane kutikule u nativnom stanju te je nađeno da se mukozni sloj, građen od kuglastih struktura, prilikom isušivanja skuplja u nepravilne nakupine. Analizom krivulja ovisnosti sile o udaljenosti dobiveni su podaci o nanomehaničkim svojstvima mukoznog sloja i same kutikule. Na temelju rezultata zaključeno je da mukozni sloj ima zaštitnu ulogu i postavljena je hipoteza prema kojoj on djeluje kao brtvilo tanjih dijelova kutikule smanjujući transpiraciju anhidrobionata.Tardigrades are microscopic invertebrates which inhabit terrestrial, freshwater and marine ecosystems. They survive adverse environmental conditions, e.g. dessication, by entering into reversible ametabolic state called cryptobiosis. Body is covered with multilaminated cuticle overlain by mucous coat. Aim of this research was to determine composition, distribution, morphology and nanomechanical properties of mucous coat on the cuticle of armoured tardigrade Echiniscus testudo using Alcian blue, colloidal iron, Nile red and Coomassie brilliant blue staining techniques, as well as Raman spectroscopy, scanning electron microscopy and atomic force microscopy. Mucous coat was found to be composed of mucopolysaccharides, lipids and proteins. It was localized mostly in dorsal epicuticle flexion zones and depressions. Atomic force microscopy imaging revealed that hydrated mucous coat, composed of globular structures, settles into an irregular aggregations upon drying. According to results obtained, protective role of mucous coat was infered; together with hypotesis according to which mucous coat acts as a sealant for thinner regions of the cuticle, thus reducing transpiration of anhydrobionts

High-throughput rat immunoglobulin G N-glycosylation profiling revealed subclass-specific changes associated with chronic stress

Immunoglobulin G (IgG) glycosylation corresponds well with immune system changes, so it can potentially be used as a biomarker for the consequences of chronic stress such as low-grade inflammation and enhanced immunosenescence in older animals. Here we present a high-throughput glycoproteomic workflow, including IgG enrichment, HILIC glycopeptide purification, and nano-LC-MS analysis of tryptic glycopeptides applied for the analysis of rat IgG. A cohort of 80 animals was exposed to seven stressors in a customized chronic stress protocol with blood and tissue sampling in three timepoints. Young female rats experienced an increase in agalactosylated glycoforms on IgG2a and IgG2c accompanied by a decrease in monogalactosylation. Among old females, increased galactosylation was observed in the IgG2b subclass, pointing to an anti-inflammatory activity of IgG. Additionally, IgG Fc N-glycosylation patterns in Sprague Dawley rats were analyzed, quantified, and reported for the first time. Our findings emphasize age-, sex- and subclass-dependent differences in IgG glycosylation related to chronic stress exposure, confirming the relevance of newly developed methods for further research in glycobiology of rodent immune response. SIGNIFICANCE: In this study, we showed that a high-throughput streamlined methodology based on protein L 96-well monolithic plates for efficient rat IgG immunoaffinity enrichment from blood plasma, paired with appropriate tryptic glycopeptide preparation, HILIC-SPE enrichment, and nano-LC-MS methods was suitable for quick processing of large sample sets. We report a subclass-specific profiling and changes in rat IgG Fc galactosylation and adrenal gland immunohistochemistry of male and female animals exposed to a customized chronic stress protocol

Distinct Longitudinal Changes in Immunoglobulin G N-Glycosylation Associate with Therapy Response in Chronic Inflammatory Diseases

Immunosuppressants and biologicals are widely used therapeutics for various chronic inflammatory diseases (CID). To gain more detailed insight into their downstream effects, we examined their impact on serum immunoglobulin G (IgG) glycosylation. We analyzed IgG subclass-specific fragment crystallizable (Fc) N-glycosylation in patients suffering from various CID using the LC-MS approach. Firstly, we compared IgG Fc N-glycosylation between 128 CID patients and 204 healthy controls. Our results replicated previously observed CID-related decrease in IgG Fc galactosylation (adjusted p-value range 1.70 × 10−2–5.95 × 10−22) and sialylation (adjusted p-value range 1.85 × 10−2–1.71 × 10−18). Secondly, to assess changes in IgG Fc N-glycosylation associated with therapy and remission status, we compared 139 CID patients receiving either azathioprine, infliximab, or vedolizumab therapy. We observed an increase in IgG Fc galactosylation (adjusted p-value range 1.98 × 10−2–1.30 × 10−15) and sialylation (adjusted p-value range 3.28 × 10−6–4.34 × 10−18) during the treatment. Furthermore, patients who reached remission displayed increased Fc galactosylation levels (p-value range 2.25 × 10−2–5.44 × 10−3) in comparison to patients with active disease. In conclusion, the alterations in IgG Fc glycosylation and the fact these changes are even more pronounced in patients who achieved remission, suggest modulation of IgG inflammatory potential associated with CID therapy

Reversal of biological age in multiple rat organs by young porcine plasma fraction

Young blood plasma is known to confer beneficial effects on various organs in mice and rats. However, it was not known whether plasma from young adult pigs rejuvenates old rat tissues at the epigenetic level; whether it alters the epigenetic clock, which is a highly accurate molecular biomarker of aging. To address this question, we developed and validated six different epigenetic clocks for rat tissues that are based on DNA methylation values derived from n = 613 tissue samples. As indicated by their respective names, the rat pan-tissue clock can be applied to DNA methylation profiles from all rat tissues, while the rat brain, liver, and blood clocks apply to the corresponding tissue types. We also developed two epigenetic clocks that apply to both human and rat tissues by adding n = 1366 human tissue samples to the training data. We employed these six rat clocks to investigate the rejuvenation effects of a porcine plasma fraction treatment in different rat tissues. The treatment more than halved the epigenetic ages of blood, heart, and liver tissue. A less pronounced, but statistically significant, rejuvenation effect could be observed in the hypothalamus. The treatment was accompanied by progressive improvement in the function of these organs as ascertained through numerous biochemical/physiological biomarkers, behavioral responses encompassing cognitive functions. An immunoglobulin G (IgG) N-glycosylation pattern shift from pro- to anti-inflammatory also indicated reversal of glycan aging. Overall, this study demonstrates that a young porcine plasma-derived treatment markedly reverses aging in rats according to epigenetic clocks, IgG glycans, and other biomarkers of aging