Cardiomyogenic differentiation of human multipotent stromal cells: a new approach fore regenerative medicine in cardiovascular disease.

Cardiovascular disease (CVD), which include hypertension, coronary heart disease (CHD), stroke, and congestive heart failure (CHF), are the leading and most common causes of mortality in both developing and industrialized countries, accounting for one third of all deaths globally. Cardiomyocyte regeneration is limited in adult life and therefore the identification of a putative source of progenitors is of great interest to provide usable in vitro models and new perspectives for regenerative therapies. This appears to be a challenge due to the peculiar functional and anatomical heart properties. The goal of this project was to differentiate adult human Adipose tissue‐derived multipotent Stem Cells (ASCs) into cardiomyocytes or their close progenitors. Thus, the ultimate aim is to develop stem cell-based therapies for the cure of cardiovascular disease. More specifically, we investigated the application of two innovative approaches: the use of human growth factors in a well defined serum-free culture medium and the application of electrical and mechanical stimuli to mimic the physiological environment of the heart. First of all, we developed a new protocol for the isolation of ASCs according with the standard GMP-procedure, which allows to extract a substantial number of cells for clinical application. The starting material was obtained from subcutaneous adipose tissue collected during an elective liposuction procedure. The isolated ASCs were characterized by flow cytometry and expanded until second passage. Several different cocktails of growth factors, cytokines and several culture conditions were tested for their ability to induce cardiomyogenesis. The goal was to identify the right combination of those factors and their time of application. One of the cardiogenic cocktails tested was indeed very effective in inducing the selected cardiac markers (e.g. NKX2-5 and MEF2C) and was chosen for the second part of this research work. The differentiation obtained was highly reproducible, as it was observed in 15 out of 15 samples from different patients. Interestingly, non induced ASCs expressed half of the selected cardiac markers (i.e. TBX5, GATA4 and BAF60C), suggesting the high Abstract V intrinsic potential and the inherent propensity of ASCs to differentiate into cardiac precursors. We further analysed the morphological changes that ASCs underwent during their differentiation by use of electron microscopy and immunofluorescence staining. By treating the cells with our selected induction cocktail, clear changes in their morphology were observed: the cells assumed a fusiform shape and became oriented. The last milestone was to understand if the combination of the cardiac induction cocktail with a mechanical (and eventually electrical) stimulation on a scaffold would increase the efficiency of cardiogenic differentiation. For this, a customized device has been developed by the Swiss Stem Cell Foundation in collaboration with Scuola Universitaria Professionale della Svizzera Italiana (SUPSI) for the application of controlled mechanical and/or electrical stimuli to a scaffold over a desired lap of time. The scaffolds used were developed and characterized by Prof. Ciardelli’s Research Group (Materials in Bionanotechnology, Department of Mechanical and Aerospace Engineering, Politecnico di Torino). The preliminary results showed that the combination of growth factors, cytokines and mechanical stimulation for 10 days led to an at least 2 fold increase in mRNA expression of NKX2‐5, MEF2C, HAND2 and MHC when compared to the induction with cardiogenic cocktail alone on a simple 2D culture condition. In conclusion, in this work we present a new protocol for the extraction of ASCs starting from adipose lipoaspirates, a new serum‐free based cardiogenic induction cocktail for ASCs and preliminary data about the differentiation in scaffold enhanced by mechanical stimulation opening a new perspective to cardiac regeneration

Cardiomyogenic differentiation of human multipotent stromal cells: a new approach fore regenerative medicine in cardiovascular disease.

Cardiovascular disease (CVD), which include hypertension, coronary heart disease (CHD), stroke, and congestive heart failure (CHF), are the leading and most common causes of mortality in both developing and industrialized countries, accounting for one third of all deaths globally. Cardiomyocyte regeneration is limited in adult life and therefore the identification of a putative source of progenitors is of great interest to provide usable in vitro models and new perspectives for regenerative therapies. This appears to be a challenge due to the peculiar functional and anatomical heart properties. The goal of this project was to differentiate adult human Adipose tissue‐derived multipotent Stem Cells (ASCs) into cardiomyocytes or their close progenitors. Thus, the ultimate aim is to develop stem cell-based therapies for the cure of cardiovascular disease. More specifically, we investigated the application of two innovative approaches: the use of human growth factors in a well defined serum-free culture medium and the application of electrical and mechanical stimuli to mimic the physiological environment of the heart. First of all, we developed a new protocol for the isolation of ASCs according with the standard GMP-procedure, which allows to extract a substantial number of cells for clinical application. The starting material was obtained from subcutaneous adipose tissue collected during an elective liposuction procedure. The isolated ASCs were characterized by flow cytometry and expanded until second passage. Several different cocktails of growth factors, cytokines and several culture conditions were tested for their ability to induce cardiomyogenesis. The goal was to identify the right combination of those factors and their time of application. One of the cardiogenic cocktails tested was indeed very effective in inducing the selected cardiac markers (e.g. NKX2-5 and MEF2C) and was chosen for the second part of this research work. The differentiation obtained was highly reproducible, as it was observed in 15 out of 15 samples from different patients. Interestingly, non induced ASCs expressed half of the selected cardiac markers (i.e. TBX5, GATA4 and BAF60C), suggesting the high Abstract V intrinsic potential and the inherent propensity of ASCs to differentiate into cardiac precursors. We further analysed the morphological changes that ASCs underwent during their differentiation by use of electron microscopy and immunofluorescence staining. By treating the cells with our selected induction cocktail, clear changes in their morphology were observed: the cells assumed a fusiform shape and became oriented. The last milestone was to understand if the combination of the cardiac induction cocktail with a mechanical (and eventually electrical) stimulation on a scaffold would increase the efficiency of cardiogenic differentiation. For this, a customized device has been developed by the Swiss Stem Cell Foundation in collaboration with Scuola Universitaria Professionale della Svizzera Italiana (SUPSI) for the application of controlled mechanical and/or electrical stimuli to a scaffold over a desired lap of time. The scaffolds used were developed and characterized by Prof. Ciardelli’s Research Group (Materials in Bionanotechnology, Department of Mechanical and Aerospace Engineering, Politecnico di Torino). The preliminary results showed that the combination of growth factors, cytokines and mechanical stimulation for 10 days led to an at least 2 fold increase in mRNA expression of NKX2‐5, MEF2C, HAND2 and MHC when compared to the induction with cardiogenic cocktail alone on a simple 2D culture condition. In conclusion, in this work we present a new protocol for the extraction of ASCs starting from adipose lipoaspirates, a new serum‐free based cardiogenic induction cocktail for ASCs and preliminary data about the differentiation in scaffold enhanced by mechanical stimulation opening a new perspective to cardiac regeneration

PROFIL SISWA DALAM MENYELESAIKAN MATERI FUNGSI KOMPOSISI DAN INVERS BERBANTU SITUS PEMBELAJARAN ONLINE

Penelitian ini bertujuan untuk melihat profil siswa dalam mengerjakan soal matematika pada materi Fungsi Komposisi dan Invers Fungsi tanpa dan dengan menggunakan situs pembelajaran online. Penelitian ini merupakan penelitian deskriptif kualitatif. Teknik pengumpulan data yang digunakan dalam penelitian ini adalah metode tes tertulis dan wawancara. Teknik analisis yang digunakan dalam penelitian ini yaitu (1) reduksi data, (2) penyajian data, dan (3) pengambilan keputusan dan verifikasi. Subjek pada penelitian ini sebanyak 5 siswa kelas X di Kota Pekalongan dengan tahun ajaran 2021/2022. Berdasarkan analisis hasil penelitian yang telah dilakukan, diketahui bahwa siswa yang mengerjakan dengan bantuan situs pembelajaran online cenderung memiliki nilai yang baik namun tidak mampu memahami proses penyelesaian tersebut. Selain itu, siswa juga cenderung untuk tidak melakukan pengecekan jawaban. Sedangkan siswa yang mengerjakan tanpa bantuan situs pembelajaran online, mengalami kesulitan dalam menyelesaikan permasalahan tersebut namun memiliki usaha untuk mencari tahu solusi melalui buku paket atau catatan saat di sekolah. Sehingga situs pembelajaran online dapat dimanfaatkan guru untuk membantu siswa dalam mengecek hasil jawaban, bukan untuk menjadi sumber dalam mencari solusi dari permasalahan yang diberikan

Triazolinedione-'clicked' poly(phosphoester)s : systematic adjustment of thermal properties

The thermal properties of halogen-free flame retardant poly(phosphoester)s from acyclic diene metathesis polycondensation have been optimized by a systematic post-modification using 1,2,4-triazoline- 3,5-dione derivatives. The straightforward modification not only increased their glass transition temperatures significantly but also improved the thermal stability with respect to their char yields

Only hearing what they want to hear: Assessing when and why performance information triggers intentions to coproduce

While performance information is often used to communicate the importance of public policies and stimulate civic engagement, we know little about the processes that connect the two. This study proposes a conceptual model that links performance information to a specific form of public engagement: coproduction. Drawing on insights from information aversion theory, we argue that the effect of performance information on engagement in coproduction depends on levels of policy understanding and the valence of performance information that individuals are exposed to. Specifically, we predict that individuals exposed to positive performance information will understand the policy better than those exposed to negative performance information. Further, we predict that higher levels of policy understanding will increase coproduction engagement intentions. These predictions are examined using two experiments and a representative sample of US residents (n = 836). Findings indicate that participants best understood positive information and that understanding significantly increased coproduction engagement intentions

Krankheitskosten, gesundheitsbezogene Lebensqualität und Versorgungssituation bei Patienten mit kognitiven Defiziten

Die vorliegende Studie befasst sich mit der Erhebung von Krankheitskosten, gesundheitsbezogener Lebensqualität und der Versorgungssituation bei Patienten mit kognitiven Defiziten mit Schwerpunkt auf der Demenz vom Alzheimertyp (DAT). Ein weiteres Ziel dieser Studie war die Identifikation von signifikanten Einflussfaktoren auf Kosten und Lebensqualität von Patienten mit DAT. Bei der vorliegenden Studie handelt es sich um eine Querschnittsstudie mit insgesamt 177 Patienten mit kognitiven Defiziten. Nach Diagnosen aufgeschlüsselt nahmen 33 Patienten teil mit „mild cognitive impairment“ (MCI), 136 Patienten mit DAT (DAT) und 8 Patienten mit einer dementiellen Mischform (MF). Zur Kostenerhebung wurden die Patienten sowie ihre betreuenden Angehörigen per Fragebogen über die aufgrund der Erkrankung entstandenen Kosten in den vergangenen drei Monaten befragt. Aus den Angaben wurden die direkten, indirekten und Gesamtkosten pro Quartal berechnet. Zudem wurden die Patienten und Angehörigen zu der aktuellen gesundheitsbezogenen Lebensqualität des Patienten befragt. Dies geschah mittels standardisierter Messinstrumente für die Dimensionen der krankheitsübergreifenden Lebensqualität (EQ-5D/EQ-VAS) als auch der krankheitsspezifischen Lebensqualität (Qol-AD). Weiterhin wurde auf das mögliche Vorliegen von einer Depression (GDS) oder neuropsychologischen Verhaltensauffälligkeiten (NPI) getestet. Zur Evaluation des Gesundheitszustandes wurden sowohl die aktuelle Kognition (MMSE, ADAS-Cog) als auch die Nebendiagnosen, der Gesamtzustand und die Alltagskompetenzen (ADCS-ADL) erhoben. Dieses umfassende Gesamtbild des Patienten ermöglichte eine in deutschen Studien bisher noch nicht durchgeführte Identifikation von Einflussfaktoren auf die Kosten und die gesundheitsbezogene Lebensqualität von Patienten mit DAT. Insgesamt ergaben sich im Rahmen dieser Studie Gesamtkosten von € 3.030 ± 3.770 pro Quartal. Eine Hochrechnung ergab jährliche Gesamtkosten von € 12.120 pro Patient. Diese setzten sich zusammen aus direkten Kosten mit einem Anteil von 75,3 % und indirekten Kosten mit einem Anteil von 24,7 %. Den größten Kostenanteil stellten hierbei die Pflegekosten mit € 1.272 ± 1.460 und damit einem Anteil von 42,0 % der Gesamtkosten dar. Die Analyse nach Einflussfaktoren ergab den größten statistischen Zusammenhang für die Fähigkeit zur Bewältigung der Aktivitäten des täglichen Lebens (p=<0,001, r=0,590) sowie der damit verbundenen Pflegebedürftigkeit der Patienten (p=<0,001, r=0,694). Die Kosten stiegen zwar auch mit zunehmenden kognitiven Defiziten, jedoch waren diese nicht der einzige ausschlaggebende Faktor für den Ressourcenverbrauch (p=<0,001, r=-0,410). Bei Patienten mit DAT zeigte sich eine eingeschränkte Lebensqualität gegenüber der Normalbevölkerung. Dabei wurde die Lebensqualität des Patienten von Angehörigen und Patienten nicht nur insgesamt unterschiedlich bewertet, sondern auch durch verschiedene Faktoren beeinflusst. Für die Patientenpopulation fand sich dabei eine bessere Eigenbewertung der Lebens¬qualität als die Fremdeinschätzung durch die pflegenden Angehörigen. Wichtig¬ster Einflussfaktor war dabei für die Patienten die Stimmungslage und für die Angehöri¬gen die Pflegebedürftigkeit sowie die Alltagsbewältigung der Patienten. Als Schlussfolgerung sollte deshalb bei der Wahl der Lebensqualität als outcome-Parameter falls möglich sowohl die Selbst- als auch die Fremdeinschätzung erhoben werden. Bei der Versorgung der Patienten zeigten sich in einigen Teilbereichen Unter- oder Fehlversorgungen. Diese betrafen insbesondere die finanzielle Versorgung (v.a. die „Demenzpauschale“) und die medika¬mentöse Versorgung. Eine bessere Informationspolitik für Patienten und Angehörige bezüglich finanzieller Unterstützung und medikamentöser Möglichkeiten scheint notwendig zu sein

Only hearing what they want to hear:Assessing when and why performance information triggers intentions to coproduce

While performance information is often used to communicate the importance of public policies and stimulate civic engagement, we know little about the processes that connect the two. This study proposes a conceptual model that links performance information to a specific form of public engagement: coproduction. Drawing on insights from information aversion theory, we argue that the effect of performance information on engagement in coproduction depends on levels of policy understanding and the valence of performance information that individuals are exposed to. Specifically, we predict that individuals exposed to positive performance information will understand the policy better than those exposed to negative performance information. Further, we predict that higher levels of policy understanding will increase coproduction engagement intentions. These predictions are examined using two experiments and a representative sample of US residents (n = 836). Findings indicate that participants best understood positive information and that understanding significantly increased coproduction engagement intentions

Rapid identification and detection of β-lactamase-producing Enterobacteriaceae from positive blood cultures by MALDI-TOF/MS.

Abstract Objectives Current evidence suggests that early diagnosis of sepsis and timely detection of antimicrobial resistance are crucial to improve mortality rates among patients. The aim of this study was to evaluate a rapid method for the identification of Gram-negative bacteria from positive blood cultures (BCs), combined with the detection of extended spectrum β-lactamases (ESβL) and carbapenemases production, by means of MALDI-TOF/MS analysis. Methods During the study, all BCs positive for Gram-negative rods were selected. Starting from bacterial pellets obtained directly from BC broths, species identification and hydrolysis assays were achieved through MALDI-TOF/MS (Bruker). In particular, we performed a hydrolysis assays of cefotaxime (CTX) and ertapenem (ERT) for the rapid detection of resistance via ESβL and carbapenemases, respectively. These results were compared with the routine workflow, including BC subcultures and confirmation phenotypic methods. Finally, a comparison of the turnaround-time (TAT) between the two protocols was conducted. Results Overall, 185 BCs positive for Enterobacteriaceae were collected. In terms of species identification, we observed a concordance of 95.9% comparing MALDI-TOF/MS results to the subculture-based method. The sensitivity and specificity for CTX hydrolysis assay were 91.1% and 92%, respectively; ERT hydrolysis assay showed a sensitivity of 96.2% and a specificity of 99.2%. The TAT of the proposed MALDI TOF/MS-based protocol was significantly lower compared with the routine workflow (P  Conclusions The proposed protocol can provide reliable bacterial identification and data concerning β-lactam resistance in only 3 hours, positively improving management of patients in terms of antimicrobial stewardship