scholarly journals Rotational Transport of Islets: The Best Way for Islets to Get around?

2013 ◽  
Vol 2013 ◽  
pp. 1-7
Author(s):  
Rupert Oberhuber ◽  
Christof Mittermair ◽  
Bettina Zelger ◽  
Daniela Pirkebner ◽  
Anna Draxl ◽  
...  
Keyword(s):  
Islet Cell ◽  
Islet Cells ◽  
Culture Conditions ◽  
Isolated Islets ◽  
Significant Difference ◽  
Glucose Tolerance Tests ◽  
Isolation Facilities ◽  
Rotary Culture

Islet transplantation is a valid treatment option for patients suffering from type 1 diabetes mellitus. To assure optimal islet cell quality, specialized islet isolation facilities have been developed. Utilization of such facilities necessitates transportation of islet cells to distant institutions for transplantation. Despite its importance, a clinically feasible solution for the transport of islets has still not been established. We here compare the functionality of isolated islets from C57BL/6 mice directly after the isolation procedure as well as after two simulated transport conditions, static versus rotation. Islet cell quality was assessed using real-time live confocal microscopy.In vivoislet function after syngeneic transplantation was determined by weight and blood sugar measurements as well as by intraperitoneal glucose tolerance tests. Vascularization of islets was documented by fluorescence microscopy and immunohistochemistry. All viability parameters documented comparable cell viability in the rotary group and the group transplanted immediately after isolation. Functional parameters assessedin vivodisplayed no significant difference between these two groups. Moreover, vascularization of islets was similar in both groups. In conclusion, rotary culture conditions allows the maintenance of highest islet quality for at least 15 h, which is comparable to that of freshly isolated islets.

In vitro lymphocyte recognition of islet cells following in vivo priming with allogeneic murine pancreatic islets

1984 ◽  
Vol 105 (1) ◽  
pp. 87-92 ◽  
Author(s):  
Joanne Scott ◽  
Peter G. MacKay ◽  
Åke Lernmark
Keyword(s):  
Lymph Node ◽  
Pancreatic Islets ◽  
Islet Cell ◽  
Islet Cells ◽  
Culture Conditions ◽  
Pancreatic Tissue ◽  
Insulin Dependent ◽  
Lymph Node Cells

Abstract. Lymphocytes from patients with insulin-dependent diabetes have been shown to be sensitized to pancreatic tissue antigens. Mice immunized with homologous pancreatic islets have been found to develop glucose intolerance and insulitis. Since lymphocytes may be involved in diabetogenesis, we wished to determine if lymph node cells from islet-immunized mice can recognize and respond to islet cells in vitro. A.TL female mice were immunized with an emulsion of BALB/c islet homogenate and complete Freund's adjuvant (CFA); sham-treated A.TL mice were injected with adjuvant and water. Mice were sacrificed 7–8 days later and the draining lymph nodes were removed. The lymph node cells were co-cultured with freshly prepared irradiated BALB/c islet cell, which served as stimulator cells. The co-cultures were incubated for 24–26 h at 37°C, followed by a 16 h [3H]thymidine (TdR) pulse. A significant proliferation of lymph node cells from islet-primed mice was induced during the in vitro stimulation with irradiated islet cells when compared with lymph node cells from sham-treated mice (P < 0.001). The response may be islet-cell-specific, since irradiated lymph node cells from BALB/c mice failed to proliferative response under the same culture conditions (P > 0.80).

scholarly journals BET Proteins Are Required for Transcriptional Activation of the Senescent Islet Cell Secretome in Type 1 Diabetes

2019 ◽  
Vol 20 (19) ◽  
pp. 4776 ◽  
Author(s):  
Peter J. Thompson ◽  
Ajit Shah ◽  
Hara Apostolopolou ◽  
Anil Bhushan
Keyword(s):  
Type 1 Diabetes ◽  
Beta Cells ◽  
Transcriptional Activation ◽  
Pancreatic Beta Cells ◽  
Islet Cells ◽  
Disease Onset ◽  
Protein Inhibition ◽  
Bet Protein

Type 1 diabetes (T1D) results from the progressive loss of pancreatic beta cells as a result of autoimmune destruction. We recently reported that during the natural history of T1D in humans and the female nonobese diabetic (NOD) mouse model, beta cells acquire a senescence-associated secretory phenotype (SASP) that is a major driver of disease onset and progression, but the mechanisms that activate SASP in beta cells were not explored. Here, we show that the SASP in islet cells is transcriptionally controlled by Bromodomain ExtraTerminal (BET) proteins, including Bromodomain containing protein 4 (BRD4). A chromatin analysis of key beta cell SASP genes in NOD islets revealed binding of BRD4 at active regulatory regions. BET protein inhibition in NOD islets diminished not only the transcriptional activation and secretion of SASP factors, but also the non-cell autonomous activity. BET protein inhibition also decreased the extent of SASP induction in human islets exposed to DNA damage. The BET protein inhibitor iBET-762 prevented diabetes in NOD mice and also attenuated SASP in islet cells in vivo. Taken together, our findings support a crucial role for BET proteins in the activation of the SASP transcriptional program in islet cells. These studies suggest avenues for preventing T1D by transcriptional inhibition of SASP.

scholarly journals Simultaneous detection of decidual Th1/Th2 and NK1/NK2 immunophenotyping in unknown recurrent miscarriage using 8-color flow cytometry with FSC/Vt extended strategy

2017 ◽  
Vol 37 (3) ◽  
Author(s):  
Peng Dong ◽  
Xi Wen ◽  
Jia Liu ◽  
Cui-Yan Yan ◽  
Jing Yuan ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Mononuclear Cells ◽  
Recurrent Miscarriage ◽  
Simultaneous Detection ◽  
Live Cells ◽  
Color Flow ◽  
Multiparametric Flow Cytometry ◽  
Significant Difference

Th1/Th2 imbalance is considered as a mechanism for recurrent miscarriage. The NK1/NK2 paradigm is hypothesised to play an important role in pregnancy. However, few results showed simultaneous changes of these subsets in vivo in decidual tissues. The present study aimed to detect the decidual mononuclear cells (dMo), and the Th1/Th2, and NK1/NK2 paradigm simultaneously using multiparametric flow cytometry (MFC) in unexplained recurrent miscarriages (URM). Mononuclear cells were isolated from the decidual tissues of URM cases and early pregnant women. The mononuclear cell percent was demonstrated by detecting the expression of CD3, CD4, CD8, CD56, and CD16 extracellular markers, interferon (IFN)-γ, and interleukin (IL)-4 intracellular markers in live cells using 8-color flow cytometry with forward scatter (FSC)/side scatter (SSC) and FSC/viability (Vt) initial gating strategies, and the ratios of Th1/Th2 and decidual NK1 (dNK1)/decidual NK2 (dNK2) cells were compared between the subject groups. Two initial gating strategies of the FSC/SSC or FSC/Vt, with central or extended gating scales, were adapted, and there was no main effect or interaction for the cell proportions, except for the type 1 and type 2 subsets in the FSC/Vt extended gating strategy. There was no significant difference of the proportions of the decidual T, dNK, NKT-like, Th, and Tc cells between the two groups. However, the Th1/Th2 and dNK1/dNK2 ratios in the URM patients were higher compared with the normal group when using the FSC/Vt extended gating strategy. The present study provides means to detect Th1/Th2 and dNK1/dNK2 simultaneously in URM patients for large sample investigations in the future.

190 EFFECT OF THE PROPERTIES OF CERVICAL MUCUS ON PREGNANCY RATES IN HOLSTEIN HEIFERS AND COWS AFTER EMBRYO TRANSFER

2006 ◽  
Vol 18 (2) ◽  
pp. 203
Author(s):  
T. Maekawa ◽  
S. Morita ◽  
O. Douchi ◽  
H. Koyama
Keyword(s):  
Embryo Transfer ◽  
Evaluation Method ◽  
Subjective Evaluation ◽  
Selection Criterion ◽  
Pregnancy Rates ◽  
Cervical Mucus ◽  
Chi Square ◽  
Significant Difference

Selection of animals as recipients of embryo transfer is an important procedure of embryo transfer on farms. Most animals are evaluated for their acceptability as recipients based on the quality of their corpus leteum (CL). However, since rectal palpation is a subjective evaluation method, a more objective method of assessing the suitability of the recipient is required. Cervical mucus may be able to be used to evaluate the condition of the uterus indirectly. The purpose of this study was to investigate the relationship between the properties of cervical mucus and pregnancy rates after embryo transfer in Holstein heifers and cows. Cervical mucus was collected using a swab off the ostium uteri externum and was stained with 5% Giemsa's solution for 20 min one day before embryo transfer. The stained cervical mucus were classified based on the type of staining pattern (Kitamura et al. 2003 Theriogenology 59, 307) into five groups: filiaceous (Type 1), taenia (Type 2), claustral (Type 3), nubecula (Type 4), or aqueous (Type 5). Proportions of the types of cervical mucus and pregnancy rates were analyzed by chi-square test. In Experiment 1, 113 heifers and 266 cows were examined for cervical mucus type. No significant difference was observed in the proportions of the types of cervical mucus between heifers and cows (heifers: 35.4%, 18.6%, 16.8%, 25.7%, and 3.5%; cows: 24.4%, 14.3%, 20.3%, 30.8%, and 10.2% for Types 1∼5, respectively). In Experiment 2, either a fresh or frozen-thawed embryo was implanted in vivo in 84 heifers and 163 cows 7 days after estrus. The heifers and cows were judged to have normal sized CLs (normal, 17 mm or more) and have no vaginal abnormalities such as cervical mucus contaminated with pus and urovagina as per vaginal examination. The proportions of acceptable Type 5 recipients was lower than that of Type 1 (P < 0.05). The pregnancy rates were 47.6% for heifers and 45.4% for cows (Table 1). The pregnancy rates of Types 1–3 (53.5%) were significantly higher than for Types 4 and 5 (29.9%) in the cows (P < 0.05). Although there was no statistically significant difference, the same tendency was observed in the heifers. Pregnancy was unsuccessful in Type 5 recipients, both heifers and cows. The total pregnancy rates of Types 1–3 were significantly higher than for Types 4 and 5 (53.5% vs. 29.9%, P < 0.001). These results suggest that cervical mucus type can serve as an objective selection criterion for embryo recipients. Further, embryo transfer should be avoided in Type 5 recipients. Table 1. Cervical mucus type and pregnancy rates (%) in dairy cattle

Measurement of pancreatic islet cell proliferation by heavy water labeling

2007 ◽  
Vol 293 (5) ◽  
pp. E1459-E1464 ◽  
Author(s):  
Songyuan Chen ◽  
Scott Turner ◽  
Ellen Tsang ◽  
Julie Stark ◽  
Holly Turner ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Heavy Water ◽  
Islet Cell ◽  
Bone Marrow Cells ◽  
Islet Cells ◽  
Therapeutic Agents ◽  
Pancreatic Islet Cell ◽  
Highly Correlated

We describe a sensitive technique for measuring long-term islet cell proliferation rates in vivo in rats. Pancreatic islets were isolated and the incorporation of deuterium (2H) from heavy water (2H2O) into the deoxyribose moiety of DNA was measured by GC-MS. The results of heavy water labeling and BrdU staining were compared. The two methods were highly correlated ( r = 0.9581, P < 0.001). Based on long-term heavy water labeling, ∼50% of islet cells divided in rats between 8 and 15 wk of age. Of interest, long-term BrdU administration suppressed proliferation of islet cells significantly, but not of bone marrow cells. Physiological evidence further supported the validity of the method: older animals (24 wk old) had 60% lower islet cell proliferation rates than younger rats (5 wk old), and partial (50%) pancreatectomy increased proliferation by 20%. In addition, cholecystokinin-8 treatment significantly stimulated proliferation in pancreatectomized rats only. In summary, heavy water labeling is a quantitative approach for measuring islet cell proliferation and testing therapeutic agents.

Overexpression of the nuclear factor-κB subunit c-Rel protects against human islet cell death in vitro

2009 ◽  
Vol 297 (5) ◽  
pp. E1067-E1077 ◽  
Author(s):  
Dariush Mokhtari ◽  
Andreea Barbu ◽  
Ilir Mehmeti ◽  
Chantal Vercamer ◽  
Nils Welsh
Keyword(s):  
Cell Death ◽  
Islet Cell ◽  
Nuclear Translocation ◽  
Islet Cells ◽  
Human Islet ◽  
Nuclear Fraction ◽  
Nuclear Factor ◽  
Subunit C

The transcription factor nuclear factor (NF)-κB is known to modulate rates of apoptosis and may therefore play a role in the increased β-cell death that occurs in type 1 and type 2 diabetes. The aim of the present investigation was to study the expression of NF-κB subunits in human islet cells and whether overexpression of the NF-κB subunit c-Rel affects islet cell survival. We detected expression of p65, Rel-B, p50, p105, p52, and the ribosomal protein S3 (rpS3) in human islet cells. Among these, only p65 and rpS3 were translocated from the cytosolic to the nuclear fraction in response to cytokines. Interestingly, rpS3 participated in p65 binding to the κB-element in gel shift analysis experiments. We observed cytoplasmic c-Rel expression in vivo in 6J mice, and signs of nuclear translocation in β-cells of infiltrated nonobese diabetic islets. Human islet cells were also dispersed by trypsin treatment and transduced with a c-Rel adenoviral vector. This resulted in increased expression of c-Rel and inhibitory factor κB, increased κB-binding activity, and augmented protein levels of Bcl-XL, c-IAP2, and heat shock protein 72. c-Rel expression in human islet cells protected against cytokine-induced caspase 3 activation and cell death. c-Rel protected also against streptozotocin- and H2O2-induced cell death, in both intact rat islets and human islet cells. We conclude that rpS3 participates in NF-κB signaling and that a genetic increase in the activity of the NF-κB subunit c-Rel results in protection against cell death in human islets.

scholarly journals What does it take to consent to islet cell xenotransplantation?: Insights from an interview study with type 1 diabetes patients and review of the literature

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Johannes Kögel ◽  
Sandra Thiersch ◽  
Barbara Ludwig ◽  
Jochen Seissler ◽  
Georg Marckmann
Keyword(s):  
Type 1 Diabetes ◽  
Informed Consent ◽  
Islet Cell ◽  
Islet Cells ◽  
Qualitative Content Analysis ◽  
Consent Form ◽  
Diabetic Patients ◽  
Porcine Islet ◽  
Informed Consent Form

Abstract Background The transplantation of porcine islet cells provides a new potential therapy to treat patients with type 1 diabetes mellitus (T1DM). Compared to other biomedical technologies, xenotransplantation stands out in terms of its involvement of animals as graft sources, as well as the possible transmission of infectious diseases. As these aspects are especially relevant for potential xenotransplantation recipients, it is important to assess their opinion regarding this technology, in particular in terms of the requirements that should be met in the informed consent process for xenotransplantation. Methods We conducted qualitative interviews with seven T1DM patients to assess their information needs prior to xenotransplantation. Before the interview, the participants received a model informed consent form for a clinical trial with porcine islet cells transplantation. The interviews were transcribed and analysed using qualitative content analysis. Results In the interviews, we identified several requirements that are crucial for patients with T1DM in order to consider xenotransplantation as a potential treatment option: therapy-related requirements, professional care and supervision, successful behaviour and attitude management, improving quality of life, and managing control/self-determination challenges. Regarding the informed consent form, several of the participants’ questions remained open and should be addressed in more detail. The interviewees stressed the importance of personal consultations. Conclusions To become a sustainable therapeutic option, patients especially expected an improved diabetes control and a reduction of diabetes-related burdens. Health-related aspects prove to be pivotal for diabetic patients when considering porcine islet cell transplantation. The use of pigs as source for organ retrievals was not considered as problematic.

188 EFFECTS OF CULTURE MEDIUM AND PROTEIN SUPPLEMENTATION ON mRNA EXPRESSION OF IN VITRO-PRODUCED PREIMPLANTATION BOVINE EMBRYOS

2008 ◽  
Vol 20 (1) ◽  
pp. 173
Author(s):  
M. N. Purpera ◽  
C. B. Ballard ◽  
A. M. Giraldo ◽  
D. Hylan ◽  
R. A. Godke ◽  
...  
Keyword(s):  
Connexin 43 ◽  
Mrna Level ◽  
Culture Conditions ◽  
Protein Supplementation ◽  
Developmental Potential ◽  
Glut 1 ◽  
Culture Systems ◽  
Significant Difference

Numerous studies have reported aberrant gene expression levels in embryos attributed to suboptimal culture conditions. This study investigated the effects of different culture systems and protein sources on the development of IVP embryos as measured by cleavage and blastocyst rates, cell number, and relative abundance levels of Oct-4, Connexin 43, Nanog, and glucose transporter-1 (Glut-1) when compared with in vivo embryos. Experiment (Exp) 1 compared IVP embryos cultured in either synthetic oviductal fluid (SOFaa) or potassium simplex optimized medium (KSOMaa) supplemented with amino acids. Exp 2 compared the same two culture systems with and without the addition of calf serum (CS). Oocytes were matured for 22 h, fertilized in vitro, and then cultured in the appropriate treatment medium. RNA was extracted from pools of blastocysts, reverse transcribed to cDNA, and primer-specific amplified via Q-PCR. Exp 1 analyzed 10 pools per treatment, and either 10 (in vivo) or 11 pools were analyzed per treatment in Exp 2. One-way ANOVA followed by multiple pair-wise comparisons using Tukey's test was used to detect differences between treatments and in vivo embryos (P < 0.05). Results from both experiments indicated that, despite similar cleavage and blastocyst rates among treatments, significant differences were detected at the mRNA level and in cell numbers between treatments. In Exp 1, Oct-4 was found to have a mean abundance mRNA level significantly greater in KSOMaa-cultured blastocysts than in either SOFaa-cultured blastocysts or in vivo embryos. The same pattern of upregulation of Oct-4 in KSOMaa or KSOMaa with CS-cultured blastocysts was detected in Exp 2. In contrast to that reported by others, Connexin 43 was not expressed at detectable levels in the in vivo embryos analyzed in our studies. Connexin 43 was not detected in IVP blastocysts used in Exp 1. Conversely, Connexin 43 was detected in KSOMaa, SOFaa, and SOFaa with CS-cultured blastocysts in Exp 2. There was no significant difference in expression of the ICM-specific transcript Nanog in either experiment. Blastocysts cultured in SOFaa with CS or KSOMaa had a significant upregulation of Glut-1 when compared with other treatments and in vivo embryos. Overall, the transcript levels of the majority of the genes analyzed were altered by the in vitro culture condition. Differences continue to be observed between in vitro-cultured and in vivo embryos, and until these differences are minimized, aberrations in in vitro development will continue to arise. Further research will possibly modify the current culture conditions, allowing for the production of in vitro embryos of higher developmental potential similar to that observed in in vivo-derived embryos.

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