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During intra-portal pancreatic islet transplantation (PITx), innate immune reactions such as the instant blood mediated inflammatory reaction (IBMIR) cause an immediate loss of islets. The non-hematopoietic erythropoietin analogue cibinetide has previously shown islet-protective effects in mouse PITx. Herein, we aimed to confirm cibinetide’s efficacy on human islets, and to characterize its effect on IBMIR. We cultured human islets with pro-inflammatory cytokines for 18 hours with or without cibinetide. ATP content and caspase 3/7 activity were measured. Dynamic glucose perfusion assay was used to evaluate islet function. To evaluate cibinetides effect on IBMIR, human islets were incubated in heparinized polyvinyl chloride tubing system with ABO compatible blood and rotated for 60 minutes to mimic the portal vein system. Moreover, human islets were transplanted into athymic mice livers via the portal vein with or without perioperative cibinetide treatment. The mice were sacrificed six days following transplantation and the livers were analyzed for human insulin and serum for human C-peptide levels. Histological examination of recipient livers to evaluate islet graft infiltration by CD11b+ cells was performed. Our results show that cibinetide maintained human islet ATP levels and reduced the caspase 3/7 activity during culture with pro-inflammatory cytokines and improved their insulin secreting capacity. In the PVC loop system, administration of cibinetide reduced the IBMIR-induced platelet consumption. In human islet to athymic mice PITx, cibinetide treatment showed an increased amount of human insulin in the livers and higher serum human C-peptide, while histological examination of the livers showed reduced infiltration of pro-inflammatory CD11b+ cells around islets grafts compared to the controls. In summary, Cibinetide protected isolated human islets in a pro-inflammatory milieu and reduced IBMIR related platelet consumption. It improved engraftment of human islets in athymic mice. The study confirms that cibinetide is a promising agent to be used in clinical PITx.

A plant fossil assemblage recently recovered from two localities belonging to the Albian Balyktakh Formation in the lower reaches of the Tuor-Yuryakh River in Kotelnyi Island (New Siberian Islands) is studied in detail. A new conifer (family Pinaceae) species Schizolepidopsis borealis Domogatskaya et Herman, sp. nov. is described. The species possesses large loose racemose polysperms (seed cones) with deeply dissected bilobate seed scales. A close association of these female cones with two species of conifer leaves, Pityophyllum ex gr. nordenskioeldii and P. ex gr. staratchinii, is reported. These three species representing reproductive and vegetative (foliar) organs probably belonged to the same whole plant species when living. Taxonomy and nomenclature of the genus Schizolepidopsis are considered and 20 new combinations are proposed and validated for previously described Schizolepis species. The palaeobotanical data for reconstructing the distribution of the genus Schizolepidopsis in space and time are based on the analysis of literature resources for 79 species covering the Late Permian to Early Cretaceous. The genus was first recorded, with some uncertainty, in the Late Permian. During the Jurassic and Early Cretaceous there was an increase in the number of Schizolepidopsis species and simultaneously the centre of the genus diversity gradually moved eastwards and north-eastwards, probably due to climate warming. It is likely that Schizolepidopsis plants became extinct close to the Albian-Cenomanian boundary due to competitive replacement by more advanced pinaceous conifers possessing compact cones with densely arranged seed scales that ensured better ovule protection.

Supplemental Material, sj-docx-1-cll-10.1177_09636897211039739 for Cibinetide Protects Isolated Human Islets in a Stressful Environment and Improves Engraftment in the Perspective of Intra Portal Islet Transplantation by Ming Yao, Anna Domogatskaya, Nils Ågren1, Masaaki Watanabe, Kazuaki Tokodai, Michael Brines, Anthony Cerami, Bo-Göran Ericzon, Makiko Kumagai-Braesch and Torbjörn Lundgren in Cell Transplantation

The criteria which are currently used for analysis and control of the investment process and the result of investing in innovative projects do not take into account all the constraints, interrelation and interdependence of processes, elements of the system and give only approximate results, confirming thus the necessity to develop new decisionmaking methods and models, best reflecting bothr actual processes and their intuitive perception. The usage of modern management practices of optimization models, based on linear programming, does not consider institutional factors, does not reflect the impact of the human factor and does not allow to reduce the riskiness due to incompleteness and a rare meeting to high-quality information. Methods of fuzzy sets theory allow to formalize not exactly formulated qualitative data, at that, by setting the value of qualitative and quantitative assessment using fuzzy numbers helps also to establish their degree of fuzziness (uncertainty), which makes it possible, based on the qualifications of the expert to use a configuration that defines the area of blur at various levels set parameters, The program of estimation of management in innovative-investment projects at the enterprise efficiency in the conditions of uncertainty and risk with use of methods of fuzzy sets theory is developed by authors in Delphi and allows to considerably simplify performing difficult calculations, to reduce calculation time, and, finally, to expand the practical use of estimation methods

After Edouard von Toll discovered Late Cretaceous plant fossils on the southern coast of New Siberia Island (Arctic Russia) in 1886 and Schmalhausen described them in 1890, no further work has been undertaken on von Toll's collection. However, palaeobotanists widely use the list of these plants based on line drawings from the paper by Schmalhausen. Here we re-examine von Toll's specimens and for the first time illustrate them by photographs. We apply а modern taxonomic nomenclature for the fossil plant remains and designate holotypes and lectotypes for the species previously described by Schmalhausen: Agathis (?) tollii (Schmalhausen) Baikovskaya, Sequoia tenuifolia (Schmalhausen) Sveshnikova et Budantsev, Nyssidium geminatum Schmalhausen and N. spicatum Schmalhausen.

The efficacy of islet transplantation for diabetes treatment suffers from lack of cadaver-derived islets, islet necrosis and long transfer times prior to transplantation. Here, we developed a method for culturing mouse and human islets in vitro on α5-laminins, which are natural components of islet basement membranes. Adhering islets spread to form layers of 1–3 cells in thickness and remained normoxic and functional for at least 7 days in culture. In contrast, spherical islets kept in suspension developed hypoxia and central necrosis within 16 h. Transplantation of 110–150 mouse islets cultured on α5-laminin-coated polydimethylsiloxane membranes for 3–7 days normalized blood glucose already within 3 days in mice with streptozotocin-induced diabetes. RNA-sequencing of isolated and cultured mouse islets provided further evidence for the adhesion and spreading achieved with α5-laminin. Our results suggest that use of such in vitro expanded islets may significantly enhance the efficacy of islet transplantation treatment for diabetes.

Need for biologically relevant coatings in regenerative medicine. Recent advances in stem cell-based regenerative medicine allow to expand human cells in almost unlimited amounts. However, expanded cells become devoid of natural niche cues and, therefore, may become prone to risks of function loss and malignant transformation. In order to maintain the expanded cells in a safe and functional way, one has to imitate in vitro the natural niche, comprising biologically relevant extracellular matrix molecules. For majority of cell types, including neurons, insulin-producing β-cells and vascular endothelial cells, biologically relevant laminins are essential part of natural niche. Laminins: 16 tissue-specific isoforms that mediate cell maintenance and behavior. Laminins (LM) are large, cross-shaped molecules that convey extracellular matrix cues via cell receptors to cell signaling systems and thus affect cell maintenance and behavior. Laminins can mediate behavior of associated cells, such as survival, adhesion, migration, proliferation, phenotype maintenance or differentiation. Each of known 16 laminin isoforms has unique biological function; mutations in laminin-encoding genes often result in severe pathologies or lethality. Despite molecular structure similarity and evolutionary homology, certain laminin isoforms may exert antagonistic effects on cell behavioral patterns. Importantly, biologically relevant laminins act in synergy with specific growth factors and cell–cell contact molecules, such as E-cadherin. Lack of either may result in malfunctioning cell culture systems. Lack of biologically relevant laminins may result in cell apoptosis, phenotype loss, or malignant transformation. Survival pathways for majority of mammalian cells depend on niche-specific extracellular matrix anchorage. Lack of such anchorage or irrelevant anchorage triggers apoptotic pathways and results in anoikis (apoptosis, caused by lack of relevant anchorage). In the absence of biologically relevant matrix cues, cells may undergo apoptosis or activate malignant pathways of anchorage-independent anti-apoptotic signaling. Majority of mammalian cell types depend on interaction with biologically relevant laminins. Biologically relevant laminins benefit quality cell culture in vitro. In vitro, cell culture systems based on use of niche-specific laminins are described for human and mouse embryonic stem cells, hematopoietic stem cells, neurons and Schwann cells, insulin-producing β-cells, and other cell types. We shall discuss success criteria and possible pitfalls for generating laminin-based safe, robust, and efficient technologies for culturing other cell types needed to treat various diseases.

Molecular phylogeneticists often find that a diversification of western Chinese plant taxa took place in the Miocene and link this to the Neogene uplift of the Tibetan Plateau. This link is made despite abundant geological evidence showing that a high but topographically complex Tibet already existed in the Paleogene. To evaluate and constrain molecular phylogenetic trees and better understand Asian plant diversification requires accurate systematic assignment of well-dated plant megafossils. Here, as part of an ongoing programme of absolute dating and taxonomic evaluation of fossil floras across southwestern China and the Himalaya, we present a new and expanded systematic treatment of the late Miocene leaf flora from the Gazhacun Formation near the village of Wang b'dui in Namling County, central southern Tibet. This flora, whose age (15 Ma) and paleoelevation (~ 5 km) are well constrained, comprises 9 families, 13 genera and 25 species (including 22 new species). The paleoflora represents a typical boreal temperate mostly deciduous broad-leaved forest attesting to a cool humid climate. At the paleolatitude of the site, < 28 °N, this is compatible with the high elevation (4700–5200 m) for this part of Tibet quantified by both leaf physiognomy and isotopic analyses. Plant fossil evidence also witnesses a much wetter Tibetan upland environment before a rising Himalaya obstructed northward-moving moist air from the Indian Ocean.

Laminins are a large family of conserved, multidomain trimeric basement membrane proteins that contribute to the structure of extracellular matrix and influence the behavior of associated cells, such as adhesion, differentiation, migration, phenotype stability, and resistance to anoikis. In lower organisms such as Hydra there is only one isoform of laminin, but higher organisms have at least 16 trimeric isoforms with varying degrees of cell/tissue specificity. In vitro protein and cell culture studies, gene manipulation in animals, and laminin gene mutations in human diseases have provided insight into the specific functions of some laminins, but the biological roles of many isoforms are still largely unexplored, mainly owing to difficulties in isolating them in pure form from tissues or cells. In this review, we elucidate the evolution of laminins, describe their molecular complexity, and explore the current knowledge of their diversity and functional aspects, including laminin-mediated signaling via membrane receptors, in vitro cell biology, and involvement in various tissues gained from animal model and human disease studies. The potential use of laminins in cell biology research and biotechnology is discussed.

How individual components of the vascular basement membrane influence endothelial cell behaviour remains unclear. Here we show that laminin α4 (Lama4) regulates tip cell numbers and vascular density by inducing endothelial Dll4/Notch signalling in vivo. Lama4 deficiency leads to reduced Dll4 expression, excessive filopodia and tip cell formation in the mouse retina, phenocopying the effects of Dll4/Notch inhibition. Lama4-mediated Dll4 expression requires a combination of integrins in vitro and integrin β1 in vivo. We conclude that appropriate laminin/integrin-induced signalling is necessary to induce physiologically functional levels of Dll4 expression and regulate branching frequency during sprouting angiogenesis in vivo.

Melanoma cells express and interact with laminins (LMs) and other basement membrane components during invasion and metastasis. In the present study we have investigated the production and migration-promoting activity of laminin isoforms in melanoma. Immunohistochemistry of melanoma specimens and immunoprecipitation/western blotting of melanoma cell lines indicated expression of laminin-111/121, laminin-211, laminin-411/421, and laminin-511/521. Laminin-332 was not detected. In functional assays, laminin-111, laminin-332, and laminin-511, but not laminin-211 and laminin-411, strongly promoted haptotactic cell migration either constitutively or following stimulation with insulin-like growth factors. Both placenta and recombinant laminin-511 preparations were highly active, and the isolated recombinant IVa domain of LMα5 also promoted cell migration. Function-blocking antibodies in cell migration assays revealed α6β1 integrin as the major receptor for laminin-111, and both α3β1 and α6β1 integrins for laminin-332 and laminin-511. In contrast, isolated LMα5 IVa domain-promoted melanoma cell migration was largely mediated via αVβ3 integrin and inhibited by RGD peptides. Given the ubiquitous expression of α5 laminins in melanoma cells and in melanoma-target tissues/anatomical structures, as well as the strong migration-promoting activity of these laminin isoforms, the α5 laminins emerge as putative primary extracellular matrix mediators of melanoma invasion and metastasis via α3β1 and other integrin receptors.

We describe a system for culturing human embryonic stem (hES) cells and induced pluripotent stem (iPS) cells on a recombinant form of human laminin-511, a component of the natural hES cell niche. The system is devoid of animal products and feeder cells and contains only one undefined component, human albumin. The hES cells self-renewed with normal karyotype for at least 4 months (20 passages), after which the cells could produce teratomas containing cell lineages of all three germ layers. When plated on laminin-511 in small clumps, hES cells spread out in a monolayer, maintaining cellular homogeneity with approximately 97% OCT4-positive cells. Adhesion of hES cells was dependent on alpha6beta1 integrin. The use of homogeneous monolayer hES or iPS cell cultures provides more controllable conditions for the design of differentiation methods. This xeno-free and feeder-free system may be useful for the development of cell lineages for therapeutic purposes.

We tested specific laminin (LN) isoforms for their ability to serve as substrata for maintaining mouse embryonic stem (ES) cells pluripotent in vitro in the absence of leukemia inhibitory factor or any other differentiation inhibitors or feeder cells. Recombinant human LN-511 alone was sufficient to enable self-renewal of mouse ES cells for up to 169 days (31 passages). Cells cultured on LN-511 maintained expression of pluripotency markers, such as Oct4, Sox2, Tert, UTF1, and Nanog, during the entire period, and cells cultured for 95 days (17 passages) were used to generate chimeric mice. LN-332 enabled ES cells proliferation but not pluripotency. In contrast, under the same conditions LN-111, Matrigel, and gelatin caused rapid differentiation, whereas LN-411 and poly-d-lysine did not support survival. ES cells formed a thin monolayer on LN-511 that differed strikingly from typical dense cluster ES cell morphology. However, expression of pluripotency markers was not affected by morphological changes. The effect was achieved at low ES cell density ( Disclosure of potential conflicts of interest is found at the end of this article.

Collagen receptor integrins alpha 1 beta 1 and alpha 2 beta 1 can selectively recognize different collagen subtypes. Here we show that their alpha I domains can discriminate between laminin isoforms as well: alpha 1I and alpha 2I recognized laminin-111, -211 and -511, whereas their binding to laminin-411 was negligible. Residue Arg-218 in alpha1 was found to be instrumental in high-avidity binding. The gain-of-function mutation E318W makes the alpha 2I domain to adopt the "open" high-affinity conformation, while the wild-type alpha 2I domain favors the "closed" low-affinity conformation. The E318W mutation markedly increased alpha 2I domain binding to the laminins (-111, -211 and -511), leading us to propose that the activation state of the alpha 2 beta 1 integrin defines its role as a laminin receptor. However, neither wild-type nor alpha 2IE318W domain could bind to laminin-411. alpha 2IE318W also bound tighter to all collagens than alpha 2I wild-type, but it showed reduced ability to discriminate between collagens I, IV and IX. The corresponding mutation, E317A, in the alpha 1I domain transformed the domain into a high-avidity binder of collagens I and IV. Thus, our results indicate that conformational activation of integrin alpha 1I and alpha 2I domains leads to high-avidity binding to otherwise disfavored collagen subtypes.

Basement membranes (BMs) consisting of laminins, collagens, and heparan sulfate proteoglycans (HSPGs) are vital for proper endothelial cell function, but many aspects of their role in vascular development remain unknown. Here, we demonstrate that vascular structures within differentiating embryoid bodies are wrapped in a BM composed of alpha4- and alpha5-chain laminins, fibronectin, collagen IV, and HSPGs. In sprouting angiogenesis, laminins were produced by stalk cells, as well as the leading tip cell, and deposited along the sprout length, including tip cell filopodia. In embryonic stem cells deficient in laminins, due to lamc1 (laminin gamma1) deletion, vascular development and organization were largely unaffected. However, the frequency of vessels with wide lumens was increased 4-fold. Laminin-deficient vessels were moreover characterized by increased fibronectin levels and enhanced endothelial cell proliferation. We conclude that laminins are dispensable for vascular development but that they regulate lumen formation in the absence of flow and vascular tone.

Extracellular matrix can influence stem cell choices, such as self-renewal, quiescence, migration, proliferation, phenotype maintenance, differentiation, or apoptosis. Three aspects of extracellular matrix were extensively studied during the last decade: physical properties, spatial presentation of adhesive epitopes, and molecular complexity. Over 15 different parameters have been shown to influence stem cell choices. Physical aspects include stiffness (or elasticity), viscoelasticity, pore size, porosity, amplitude and frequency of static and dynamic deformations applied to the matrix. Spatial aspects include scaffold dimensionality (2D or 3D) and thickness; cell polarity; area, shape, and microscale topography of cell adhesion surface; epitope concentration, epitope clustering characteristics (number of epitopes per cluster, spacing between epitopes within cluster, spacing between separate clusters, cluster patterns, and level of disorder in epitope arrangement), and nanotopography. Biochemical characteristics of natural extracellular matrix molecules regard diversity and structural complexity of matrix molecules, affinity and specificity of epitope interaction with cell receptors, role of non-affinity domains, complexity of supramolecular organization, and co-signaling by growth factors or matrix epitopes. Synergy between several matrix aspects enables stem cells to retain their function in vivo and may be a key to generation of long-term, robust, and effective in vitro stem cell culture systems.

Extracellular matrix can influence stem cell choices, such as self-renewal, quiescence, migration, proliferation, phenotype maintenance, differentiation, or apoptosis. Three aspects of extracellular matrix were extensively studied during the last decade: physical properties, spatial presentation of adhesive epitopes, and molecular complexity. Over 15 different parameters have been shown to influence stem cell choices. Physical aspects include stiffness (or elasticity), viscoelasticity, pore size, porosity, amplitude and frequency of static and dynamic deformations applied to the matrix. Spatial aspects include scaffold dimensionality (2D or 3D) and thickness; cell polarity; area, shape, and microscale topography of cell adhesion surface; epitope concentration, epitope clustering characteristics (number of epitopes per cluster, spacing between epitopes within cluster, spacing between separate clusters, cluster patterns, and level of disorder in epitope arrangement), and nanotopography. Biochemical characteristics of natural extracellular matrix molecules regard diversity and structural complexity of matrix molecules, affinity and specificity of epitope interaction with cell receptors, role of non-affinity domains, complexity of supramolecular organization, and co-signaling by growth factors or matrix epitopes. Synergy between several matrix aspects enables stem cells to retain their function in vivo and may be a key to generation of long-term, robust, and effective in vitro stem cell culture systems.

Endocrine pancreatic beta cells require endothelial signals for their differentiation and function. However, the molecular basis for such signals remains unknown. Here, we show that beta cells, in contrast to the exocrine pancreatic cells, do not form a basement membrane. Instead, by using VEGF-A, they attract endothelial cells, which form capillaries with a vascular basement membrane next to the beta cells. We have identified laminins, among other vascular basement membrane proteins, as endothelial signals, which promote insulin gene expression and proliferation in beta cells. We further demonstrate that beta1-integrin is required for the beta cell response to the laminins. The proposed mechanism explains why beta cells must interact with endothelial cells, and it may apply to other cellular processes in which endothelial signals are required.

The Schwann cell basal lamina (BL) is required for normal myelination. Loss or mutations of BL constituents, such as laminin-2 (α2β1γ1), lead to severe neuropathic diseases affecting peripheral nerves. The function of the second known laminin present in Schwann cell BL, laminin-8 (α4β1γ1), is so far unknown. Here we show that absence of the laminin α4 chain, which distinguishes laminin-8 from laminin-2, leads to a disturbance in radial sorting, impaired myelination, and signs of ataxia and proprioceptive disturbances, whereas the axonal regenerative capacity is not influenced.In vitrostudies show poor axon growth of spinal motoneurons on laminin-8, whereas it is extensive on laminin-2. Schwann cells, however, extend longer processes on laminin-8 than on laminin-2, and, in contrast to the interaction with laminin-2, solely use the integrin receptor α6β1 in their interaction with laminin-8. Thus, laminin-2 and laminin-8 have different critical functions in peripheral nerves, mediated by different integrin receptors.

The ability of a human artery to pass through 150 million liters of blood sustaining 2 billion pulsations of blood pressure with minor deterioration depends on unique construction of the arterial wall. Viscoelastic properties of this construction enable to re-seal the occuring damages apparently without direct immediate participance of the constituent cells. Collagen structures are considered to be the elements that determine the mechanoelastic properties of the wall in parallel with elastin responsible for elasticity and resilience. Collagen scaffold architecture is the function-dependent dynamic arrangement of a dozen different collagen types composing three distinct interacting forms inside the extracellular matrix of the wall. Tightly packed molecules of collagen types I, III, V provide high tensile strength along collagen fibrils but toughness of the collagen scaffold as a whole depends on molecular bonds between distinct fibrils. Apart of other macromolecules in the extracellular matrix (ECM), collagen-specific interlinks involve microfilaments of collagen type VI, meshwork-organized collagen type VIII, and FACIT collagen type XIV. Basement membrane collagen types IV, XV, XVIII and cell-associated collagen XIII enable transmission of mechanical signals between cells and whole artery matrix. Collagen scaffold undergoes continuous remodeling by decomposition promoted with MMPs and reconstitution from newly produced collagen molecules. Pulsatile stress-strain load modulates both collagen synthesis and MMP-dependent collagen degradation. In this way the ECM structure becomes adoptive to mechanical challenges. The mechanoelastic properties of the arterial wall are changed in atherosclerosis concomitantly with collagen turnover both type-specific and dependent on the structure. Improving the feedback could be another approach to restore sufficient blood circulation.

Lack of robust methods for establishment and expansion of pluripotent human embryonic stem (hES) cells still hampers development of cell therapy. Laminins (LN) are a family of highly cell-type specific basement membrane proteins important for cell adhesion, differentiation, migration and phenotype stability. Here we produce and isolate a human recombinant LN-521 isoform and develop a cell culture matrix containing LN-521 and E-cadherin, which both localize to stem cell niches in vivo. This matrix allows clonal derivation, clonal survival and long-term self-renewal of hES cells under completely chemically defined and xeno-free conditions without ROCK inhibitors. Neither LN-521 nor E-cadherin alone enable clonal survival of hES cells. The LN-521/E-cadherin matrix allows hES cell line derivation from blastocyst inner cell mass and single blastomere cells without a need to destroy the embryo. This method can facilitate the generation of hES cell lines for development of different cell types for regenerative medicine purposes.

The establishment of stable recombinant protein-producing mammalian cell lines is an expensive, time-consuming, tedious procedure. In some cases, expressed recombinant proteins have adverse effects on host cell function, including cell adhesion. Based on the adhesive properties of SCARA5, a scavenger receptor (SR) of the class A SR family, we developed a method for selection of stable recombinant protein-producing cell clones that relies on an internal ribosome entry site (IRES) vector where the protein of interest is expressed in the same messenger RNA as SCARA5, resulting in improved adhesion and increased cell viability of recombinant protein-producing cells in serum-free media. This method does not depend on antibiotics, complicated selective cell culture media or equipment, and thus offers the advantages of being inexpensive, environmentally friendly, and simple.