Polyhydroxybutyrate-based osteoinductive mineralized electrospun structures that mimic components and tissue interfaces of the osteon for bone tissue engineering.

Scaffolds for bone tissue engineering should enable regeneration of bone tissues with its native hierarchically organized extracellular matrix (ECM) and multiple tissue interfaces. To achieve this, inspired by the structure and properties of bone osteon, we fabricated polyhydroxybutyrate (PHB)-based mineralized electrospun fibrous scaffolds. After studying multiple PHB-based fibers, we chose 7%PHB/1%Gelatin fibers (PG) to fabricate mineralized fibers that mimic mineralized collagen fibers in bone. The mineralized PG (mPG) surface had a rough, hydrophilic layer of low crystalline calcium phosphate which was biocompatible to bone marrow stromal cells (BMSCs), induced their proliferation and was osteoinductive. Subsequently, by modulating the electrospinning process, we fabricated mPG-based novel higher order fibrous scaffolds that mimic the macroscale geometries of osteons of bone ECM. Inspired by the aligned collagen fibers in bone lamellae, we fabricated mPG scaffolds with aligned fibers that could direct anisotropic elongation of mouse BMSC (mBMSCs). Further, we fabricated electrospun mPG-based osteoinductive tubular constructs which can mimic cylindrical bone components like osteons or lamellae or be used as long bone analogues based on their dimensions. Finally, to regenerate tissue interfaces in bone, we introduced a novel bi-layered scaffold-based approach. An electrospun bi-layered tubular construct that had PG in the outer layer and 7%PHB/0.5%Polypyrrole fibers (PPy) in the inner layer was fabricated. The bi-layered tubular construct underwent preferential surface mineralization only on its outer layer. This outer mineralized layer supported osteogenesis while the inner PPy layer could support neural cell growth. Thus, the bi-layered tubular construct may be used to regenerate haversian canal in the osteons which hosts nerve fibers. Overall, the study introduced novel techniques to fabricate biomimetic structures that can regenerate components of bone osteon and its multiple tissue interfaces. The study lays foundation for the fabrication of a modular scaffold that can regenerate bone with its hierarchical structure and complex tissue interfaces.

Directing ligament-mimetic bi-directional cell organization in scaffolds through zone-specific microarchitecture for ligament tissue engineering.

Ligament tissues exhibit zone-specific anisotropic cell organization. The cells in ligament-proper are longitudinally oriented, whereas, the cells in epiligament are circumferentially oriented. Therefore, scaffolds developed to regenerate ligament tissues should possess adequate architectural features to govern ligament-mimetic bi-directional cell organization. The scaffold architectural features along with ligament-mimetic cell organization may ultimately yield neo-tissues with ligament-like extracellular matrix (ECM) structure and biomechanical properties. Towards this goal, we fabricated a silk/gelatin-based core-shell scaffold (csSG) with zone-specific anisotropic architectural features, wherein, the core of the scaffold possessed longitudinally aligned pores while the shell of the scaffold possessed parallel microgrooves that are aligned circumferentially around the surface of the scaffold. The ligament-mimetic architectural features significantly improved the mechanical properties of the scaffold. Moreover, architectural features of the csSG scaffold governed zone-specific anisotropic organization of cells. The cells in the core were longitudinally oriented as observed in the ligament-proper and the cells on the shell were circumferentially oriented as observed in epiligament. This bi-directional cell orientation partially mimicked the complex cellular network in native ligament tissue. Additionally, both the core and the shell individually supported fibrogenic differentiation of stem cells which further improved their potential for ligament tissue engineering. Further, the aligned pores of the core could govern unidirectional organization of ECM deposited by cells which is crucial for regenerating anisotropic tissues like ligaments. Finally, when implanted subcutaneously in mice, the scaffolds retained their anisotropic architecture for at least 2 weeks, were biocompatible, supported cell infiltration and governed anisotropic organization of cells and ECM. Taken together, the fabricated biomimetic csSG scaffold, through its zone-specific architectural features, could govern ligament-mimetic cellular and ECM organization which is ultimately expected to achieve regeneration of ligament tissues with native-like hierarchical structure and biomechanical properties. Consequently, this study introduces bi-directional structural parameters as design criteria for developing scaffolds for ligament tissue engineering.

Quality of life among those with diabetes mellitus type II attending a secondary outpatient facility in South India.

Context and Aim: Diabetes mellitus (DM) can affect one's quality of life (QoL). Literature on the association of QoL among type II diabetics with drug compliance and diet quality among rural communities is poor. This study aimed to determine the QoL among those with type II DM attending an outpatient clinic at a secondary hospital in Tamil Nadu. Settings and Design: A cross-sectional, interview-based study was carried out among those with type II DM. A questionnaire comprising the WHO-BREF tool, Diabetes Healthy Eating Index, Hill-Bone Medication Adherence Scale was administered to participants selected via systematic random sampling. Results: The prevalence of good QoL was estimated to be 51.7% (n = 45, 95% CI: 41.20-62.20). There was no association between good QoL and medication compliance. None of the patients had a good diet quality. Bivariate analysis revealed significant association (p < 0.05) between good QoL and higher education (OR-2.70), those not on medication for complications (OR-2.81) and decreased frequency of general random blood sugar (GRBS) monitoring (OR-2.44). Multivariable analysis adjusting for gender, education, treatment/medication for complications, hospitalisation for DM and GRBS frequency demonstrated significant association between good QoL, lack of medication for complications/co-morbidities and decreased GRBS monitoring frequency with likelihood ratios of 3.25 and 3.44, respectively. Conclusions: The drop in QoL observed could be due to the aftermath of the COVID pandemic. Keeping in mind the impact of healthcare interventions on the QoL of type II DM patients, primary physicians must consider dietary and treatment plans suited to their socio-economic status.

Sulfated carboxymethylcellulose conjugated electrospun fibers as a growth factor presenting system for tissue engineering.

Inspired by the natural electrostatic interaction of cationic growth factors with anionic sulfated glycosaminoglycans in the extracellular matrix, we developed electrospun poly(hydroxybutyrate)/gelatin (PG) fibers conjugated with anionic sulfated carboxymethylcellulose (sCMC) to enable growth factor immobilization via electrostatic interaction for tissue engineering. The fibrous scaffold bound cationic molecules, was cytocompatible and exhibited a remarkable morphological and functional stability. Transforming growth factor-ß1 immobilized on the sCMC conjugated fibers was retained for at least 4 weeks with negligible release (3%). Immobilized fibroblast growth factor-2 and connective tissue growth factor were bioactive and induced proliferation and fibrogenic differentiation of infrapatellar fat pad derived mesenchymal stem cells respectively with efficiency similar to or better than free growth factors. Taken together, our studies demonstrate that sCMC conjugated PG fibers can immobilize and retain function of cationic growth factors and hence show potential for use in various tissue engineering applications.

Understanding the role of m-health to improve well-being in spouses of patients with bipolar disorder.

OBJECTIVES: Spouses and partners of individuals with bipolar disorder (BD) experience significant burden. As there are some limitations to standard psychosocial caregiver interventions, mobile health technology (mHealth) may be a way to reduce burden and improve well-being in these caregivers. The purpose of this study was to explore how the well-being of spouses or partners of patients with BD can be improved through mHealth technology. METHODS: Using a qualitative design, we conducted five focus groups and one in-depth individual interview to collect information from participants about what they would expect from such a device. The sample consists of thirteen participants (eleven spouses and two partners). The age range was 29-65, with eight females and five males. Data were collected using minimally structured interviews and independently analyzed by the authors using content analysis. RESULTS: Results indicated that the mHealth device many be helpful in at least six areas: reduction of stressors, decreased social isolation, improving communication in the relationship between the spouses, speaking with children about the illness, managing medications, and providing information on resources. CONCLUSION: Mobile health technology may be a feasible, available, and cost-effective support tool for spouses and partners of individuals with BD, especially in reducing caregiver stress. Future research is needed to develop the application and test its effectiveness on health outcomes in a larger trial.

Co-culture of infrapatellar fat pad-derived mesenchymal stromal cells and articular chondrocytes in plasma clot for cartilage tissue engineering.

BACKGROUND: Cell source plays a deterministic role in defining the outcome of a cell-based cartilage regenerative therapy and its clinical translational ability. Recent efforts in the direction of co-culture of two or more cell types attempt to combine the advantages of constituent cell types and negate their demerits. METHODS: We examined the potential of co-culture of infrapatellar fat pad-derived mesenchymal stromal cells (IFP MSCs) and articular chondrocytes (ACs) in plasma clots in terms of their ratios and culture formats for cartilage tissue engineering. RESULTS AND DISCUSSION: It was observed that IFP MSCs and ACs interact positively to produce a better quality hyaline cartilage-like matrix. While a supra-additive deposition of sulfated Glycosaminoglycans (sGAG), collagen type II, aggrecan and link protein was observed, deposition of collagen type I and X was sub-additive. (Immuno)-histologically similar cartilage was generated in vitro in IFP MSC:AC ratio of 50:50 and pure AC groups thus yielding a hyaline cartilage with 50% reduced requirement of ACs. Subsequently, we investigated if this response could be improved further by enabling better cell-cell interactions using scaffold-free systems such as self-assembled cartilage or by encapsulating cellular micro-aggregates in plasma clot. However, it was inferred that while self-assembly may have enabled better cell-cell interaction, poor cell survival negated its overall beneficial role, whereas the micro-aggregate group demonstrated highly heterogeneous matrix deposition within the construct, thus diminishing its translational utility. Overall, it was concluded that co-culture of IFP MSCs and ACs at a ratio of 50:50 within plasma clots demonstrated potential for cell-based cartilage regenerative therapy.

Scaffolds containing chitosan/carboxymethyl cellulose/mesoporous wollastonite for bone tissue engineering.

Scaffold based bone tissue engineering utilizes a variety of biopolymers in different combinations aiming to deliver optimal properties required for bone regeneration. In the current study, we fabricated bio-composite scaffolds containing chitosan (CS), carboxymethylcellulose (CMC) with varied concentrations of mesoporous wollastonite (m-WS) particles by the freeze drying method. The CS/CMC/m-WS scaffolds were characterized by the SEM, EDS and FT-IR studies. Addition of m-WS particles had no effect on altering the porosity of the scaffolds. m-WS particles at 0.5% concentration in the CS/CMC scaffolds showed significant improvement in the bio-mineralization and protein adsorption properties. Addition of m-WS particles in the CS/CMC scaffolds significantly reduced their swelling and degradation properties. The CS/CMC/m-WS scaffolds also showed cyto-friendly nature to human osteoblastic cells. The osteogenic potential of CS/CMC/m-WS scaffolds was confirmed by calcium deposition and expression of an osteoblast specific microRNA, pre-mir-15b. Thus, the current investigations support the use of CS/CMC/m-WS scaffolds for bone tissue engineering applications.

Biomaterials mediated microRNA delivery for bone tissue engineering.

Bone tissue engineering is an alternative strategy to overcome the problems associated with traditional treatments for bone defects. A number of bioactive materials along with new techniques like porous scaffold implantation, gene delivery, 3D organ printing are now-a-days emerging for traditional bone grafts and metal implants. Studying the molecular mechanisms through which these biomaterials induce osteogenesis is an equally hot field. Biomaterials could determine the fate of a cell via microRNAs (miRNAs). miRNAs are short non-coding RNAs that act as post-transcriptional regulators of gene expression and play an essential role for regulation of cell specific lineages including osteogenesis. Thus, this review focuses the recent trends on establishing a link of biomaterials with miRNAs and their delivery for bone tissue engineering applications.

A propagating ERKII switch forms zones of elevated dendritic activation correlated with plasticity.

Strong inputs to neurons trigger complex biochemical events leading to synaptic plasticity. These biochemical events occur at many spatial scales, ranging from submicron dendritic spines to signals that propagate hundreds of microns from dendrites to the nucleus. ERKII is an important signaling molecule that is involved in many aspects of plasticity, including local excitability, communication with the nucleus, and control of local protein synthesis. We observed that ERKII activation spreads long distances in apical dendrites of stimulated hippocampal CA1 pyramidal neurons. We combined experiments and models to show that this >100 mum spread was too large to be explained by biochemical reaction-diffusion effects. We show that two modes of calcium entry along the dendrite contribute to the extensive activation of ERKII. We predict the occurrence of feedback between biophysical events resulting in calcium entry, and biochemical events resulting in ERKII activation. This feedback causes a switch-like propagation of ERKII activation, coupled with enhanced electrical excitability, along the apical dendrite. We propose that this propagating switch forms zones on dendrites in which plasticity is facilitated.

Synaptic plasticity in vitro and in silico: insights into an intracellular signaling maze.

Synaptic plasticity provides a record of neuronal activity and is a likely basis for memory. The early apparent simplicity of the process of synaptic plasticity has been lost in a flood of experimental data that now implicates some 200 signaling molecules in cellular memory. It is now clear that these signaling networks perform surprisingly sophisticated cellular decisions that weigh factors such as input patterns, location of stimulus, history of activity, and context. Computer models have followed experiments into this maze of molecular detail, often matching closely with their experimental counterparts, but perhaps losing simplicity in the process. Here, we suggest that the merger of models and experiment have begun to restore the earlier simplicity by outlining a few key functional roles for signaling networks in synaptic plasticity. In this review, we discuss the current state of understanding of synaptic plasticity in terms of models and experiments.

A role for ERKII in synaptic pattern selectivity on the time-scale of minutes.

Stimulus reinforcement strengthens learning. Intervals between reinforcement affect both the kind of learning that occurs and the amount of learning. Stimuli spaced by a few minutes result in more effective learning than when massed together. There are several synaptic correlates of repeated stimuli, such as different kinds of plasticity and the amplitude of synaptic change. Here we study the role of signalling pathways in the synapse on this selectivity for spaced stimuli. Using the in vitro hippocampal slice technique we monitored long-term potentiation (LTP) amplitude in CA1 for repeated 100-Hz, 1-s tetani. We observe the highest LTP levels when the inter-tetanus interval is 5-10 min. We tested biochemical activity in the slice following the same stimuli, and found that extracellular signal-regulated kinase type II (ERKII) but not CaMKII exhibits a peak at about 10 min. When calcium influx into the slice is buffered using AM-ester calcium dyes, amplitude of the physiological and biochemical response is reduced, but the timing is not shifted. We have previously used computer simulations of synaptic signalling to predict such temporal tuning from signalling pathways. In the current study we consider feedback and feedforward models that exhibit temporal tuning consistent with our experiments. We find that a model incorporating post-stimulus build-up of PKM zeta acting upstream of mitogen-activated protein kinase is sufficient to explain the observed temporal tuning. On the basis of these combined experimental and modelling results we propose that the dynamics of PKM activation and ERKII signalling may provide a mechanism for functionally important forms of synaptic pattern selectivity.

Modelling protein functional domains in signal transduction using Maude.

Modelling of protein-protein interactions in signal transduction is receiving increased attention in computational biology. This paper describes recent research in the application of Maude, a symbolic language founded on rewriting logic, to the modelling of functional domains within signalling proteins. Protein functional domains (PFDs) are a critical focus of modern signal transduction research. In general, Maude models can simulate biological signalling networks and produce specific testable hypotheses at various levels of abstraction. Developing symbolic models of signalling proteins containing functional domains is important because of the potential to generate analyses of complex signalling networks based on structure-function relationships.

Global geometric indicator of chaos and Lyapunov exponents in Hamiltonian systems.

Recently a geometric description of chaos in Hamiltonian systems has been formulated using the tools of Riemannian geometry. Here, Hamiltonian chaos is explained in terms of the curvature properties of the configuration space manifold. In particular, it has been claimed that the average of an appropriately defined sectional curvature (K((2))) over a constant energy manifold is a measure of the global extent of chaoticity for systems with a small number of degrees of freedom. We investigate the relations between this quantity K((2)) and the maximal Lyapunov exponent lambda for some Hamiltonian systems of physical interest with two degrees of freedom. We find that there is a close relation between K((2)) and lambda(2). Both the quantities scale as E(1/2) for quartic potentials, where E is the energy. They are expected to scale as E((n-2)/n) for a general potential of degree n. However, we find that though K((2)) is a global indicator of chaos, it is not a sufficiently accurate measure of order-chaos transitions, in all cases.

Cloning, expression, and crystallization of Cpn60 proteins from Thermococcus litoralis.

Two genes of the extreme thermophilic archaeon Thermococcus litoralis homologous to those that code for Cpn60 chaperonins were cloned and expressed in Escherichia coli. Each of the Cpn60 subunits as well as the entire Cpn60 complex crystallize in a variety of morphological forms. The best crystals diffract to 3.6 A resolution at room temperature and belong to the space group 1422 with unit cell parameters a = b = 193.5 A, c = 204.2 A.

Spontaneous perforation of the common bile duct in infancy: role of Tc-99m mebrofenin hepatobiliary imaging.

A 2-month-old infant had jaundice that began 3 days after birth. The clinical features were suggestive of biliary obstruction. Tc-99m mebrofenin hepatobiliary imaging confirmed the diagnosis of spontaneous perforation of the common bile duct, and surgical intervention resulted in progressive recovery.

Alternative algorithm for the computation of Lyapunov spectra of dynamical systems.

Recently a new method for the computation of Lyapunov exponents that does not require rescaling and reorthogonalization was proposed [Rangarajan, Habib, and Ryne, Phys. Rev. Lett. 80, 3747 (1998)]. In this paper we make a detailed numerical comparison of the new method and a standard algorithm, as regards accuracy and efficiency, by applying them to some typical two-, three-, and four-dimensional systems. We find that in most cases there is reasonable agreement between the Lyapunov spectra obtained using the two algorithms. The CPU times required for computation are also comparable. However, in certain strongly chaotic cases, the new method was found to be either inefficient (taking a lot of CPU time for computation) or inaccurate.

Human Hsp70 molecular chaperone binds two calcium ions within the ATPase domain.

BACKGROUND: The 70 kDa heat shock proteins (Hsp70) are a family of molecular chaperones, which promote protein folding and participate in many cellular functions. The Hsp70 chaperones are composed of two major domains. The N-terminal ATPase domain binds to and hydrolyzes ATP, whereas the C-terminal domain is required for polypeptide binding. Cooperation of both domains is needed for protein folding. The crystal structure of bovine Hsc70 ATPase domain (bATPase) has been determined and, more recently, the crystal structure of the peptide-binding domain of a related chaperone, DnaK, in complex with peptide substrate has been obtained. The molecular chaperone activity and conformational switch are functionally linked with ATP hydrolysis. A high-resolution structure of the ATPase domain is required to provide an understanding of the mechanism of ATP hydrolysis and how it affects communication between C- and N-terminal domains. RESULTS: The crystal structure of the human Hsp70 ATPase domain (hATPase) has been determined and refined at 1. 84 A, using synchrotron radiation at 120K. Two calcium sites were identified: the first calcium binds within the catalytic pocket, bridging ADP and inorganic phosphate, and the second calcium is tightly coordinated on the protein surface by Glu231, Asp232 and the carbonyl of His227. Overall, the structure of hATPase is similar to bATPase. Differences between them are found in the loops, the sites of amino acid substitution and the calcium-binding sites. Human Hsp70 chaperone is phosphorylated in vitro in the presence of divalent ions, calcium being the most effective. CONCLUSIONS: The structural similarity of hATPase and bATPase and the sequence similarity within the Hsp70 chaperone family suggest a universal mechanism of ATP hydrolysis among all Hsp70 molecular chaperones. Two calcium ions have been found in the hATPase structure. One corresponds to the magnesium site in bATPase and appears to be important for ATP hydrolysis and in vitro phosphorylation. Local changes in protein structure as a result of calcium binding may facilitate phosphorylation. A small, but significant, movement of metal ions and sidechains could position catalytically important threonine residues for phosphorylation. The second calcium site represents a new calcium-binding motif that can play a role in the stabilization of protein structure. We discuss how the information about catalytic events in the active site could be transmitted to the peptide-binding domain.