Pancreatic cancer 3D cell line organoids (CLOs) maintain the phenotypic characteristics of organoids and accurately reflect the cellular architecture and heterogeneity In vivo

Pancreatic cancer is a highly lethal disease. Therapeutic resistance to chemotherapy is a major cause of treatment failure and recurrence in pancreatic cancer. Organoids derived from cancer stem cells (CSC) are promising models for the advancement of personalised therapeutic responses to inform clinical decisions. However, scaling-up of 3D organoids for high-throughput screening is time-consuming and costly. Here, we successfully developed organoid-derived cell lines (2.5D) from 3D organoids; the cells were then expanded and recapitulated back into organoids known as cell line organoids (CLOs). The 2.5D lines were cultured long term into 2D established cell lines for downstream comparison analysis. Experimental characterisation of the models revealed that the proliferation of CLOs was slightly faster than that of parental organoids. The therapeutic response to chemotherapeutic agents in 3D CLOs and organoids showed a similar responsive profile. Compared to 3D CLOs and organoids, 2D cell lines tended to be less responsive to all the drugs tested. Stem cell marker expression was higher in either 3D CLOs or organoids compared to 2D cell lines. An in vivo tumorigenicity study found CLOs form tumours at a similar rate to organoids and retain enhanced CSC marker expression, indicating the plasticity of CSCs within the in vivo microenvironment

Development of Potent, Conformation-Specific Inhibitors of Connexin-Mediated Communication

Connexin (Cx) mimetic peptides derived from extracellular loop II (ECL2) sequences of Cx43 (e.g. Gap 27: SRPTEKTIFII; Peptide5: VDCFLSRPTEKT) have been used as reversible, Cx-specific blockers of hemichannel (HCh) and gap junction channel (GJCh) function. These blockers typically require high concentrations (HCh: ~5 µM, 1 hour) to achieve inhibition. We report here that addition of a hexadecyl (Hdc) lipid moiety to the conserved SRPTEKT peptide sequence, SRPTEKT-Hdc, results in a novel, highly efficacious and potent inhibitor of mechanically-induced Ca2+-wave propagation (IC50: 64.8 pM) and HCh-mediated dye uptake (IC50: 45.0 pM) in Madin-Darby Canine Kidney cells expressing mCx43 (MDCK43). The lack of similar effect on dye coupling suggested channel conformation-specific inhibition, and indeed, SRPTEKT-Hdc inhibition of Ca2+-wave propagation and dye uptake was selective for Cx43 channels in the functional configuration governed by phosphorylation at serine S368 (pS368). As such, SRPTEKT-Hdc potently inhibited Ca2+-wave propagation (IC50: 230.4 pM) and dye uptake in MDCK cells expressing single site mutants of Cx43 that mimicked (MDCK43-S368D) or favored (MDCK43-S365A) phosphorylation at S368. In contrast, Ca2+-wave propagation and dye uptake were largely unaffected (IC50: 12.3 μM) by SRPTEKT-Hdc in MDCK43-S368A and -S365D cells, mutations that mimic or favor dephosphorylation at S368. Finally, we preliminarily report SRPTEKT-Hdc inhibition of Cx37-mediated Ca2+-wave propagation and dye uptake in MDCK cells expressing a doxycycline-inducible form of mCx37 (iMDCK37). Inhibition occurred in a manner similar to that observed for wild-type Cx43 channels, highlighting the likely involvement of the identical SRPTEKT sequence shared between ECL2 domains of these two Cx isotypes. Together, these data indicate that SRPTEKT-Hdc is a potent and specific inhibitor of the pS368 phospho-form of Cx43 with potential for cross-isotype inhibition of other SRPTEKT-containing Cx isotypes (possibly also in a phosphorylation-dependent manner)

Occupy feminism

The Occupy movement galvanized people from across the United States and around the world to stand up against corporate profiteering and political corruption. The revolutionary energy spread from community to community and Occupiers began to create a reality moving beyond racism, hierarchy, and patriarchy. Feminist activists, both women and men, had a major voice in Occupy. They called into question sexist behavior, unrecognized privilege and gender inequality in conjunction with other forms of oppression. Unfortunately the breakdown of the encampments and the backlash in the mainstream media led to a critical view of the movement. In reality, Occupiers formed networks of mutual aid which continue to expand and transform today. Based on ethnographic research conducted at Occupy Los Angeles and Occupy Long Beach, this thesis project explores a feminist perspective of the Occupy movement. Inspired by applied visual anthropology and new media, Occupy Feminism is an interactive zine interspersed with writing, photographs and videos. This project is an educational resource for those who want to understand feminist theory, the Occupy movement and how they intersect

How the Antiwar Movement Views the Islamic Faith

I want to explore how the antiwar movement on the University of Illinois campus views the Islamic faith and Muslims as both religious and cultural group. I want to see if their views changed post 9/11 or if they were already informed about Islam. I also want to explore why there are so few Muslims that participate in groups such as the Campus Antiwar Network (CAN), a national student organization with a chapter on the U of I campus, and Anti-War Anti-Racism Effort (AWARE), a local community group. Since this is a native study, as I am a member of CAN, I plan to observe certain times, such as meetings, rallies, awareness events, specifically for my research. I plan to use interviews and participant observation as my main tactics of research. I want to explore the language they use while referring to Muslims and see if there is a correlation in their mind to faith and terrorism

CD82 Regulation of Hematopoietic Stem/Progenitor Cell - Niche Adhesion

The spatial organization and dynamics of proteins and lipids within the cell membrane is important for the regulation of cell signaling, adhesion, and cell communication. Within the bone marrow niche, communication between hematopoietic stem/progenitor cells (HSPCs) and niche cells is essential for regulating their proliferation, differentiation, and survival. Our previous work has ascertained that HSPCs utilize a polarized domain on the plasma membrane that serves as the contact site with osteoblasts, which are important members of the bone marrow niche. Using human primary CD34+ stem/progenitor cells and the progenitor-like KG1a cell line, we found this domain to be enriched in the specific tetraspanin proteins, CD63, CD81, and CD82. Tetraspanins are multi-spanning membrane proteins that act as scaffolds for the organization of membrane domains important for regulating adhesion and signaling. CD82 is of particular interest, as it is highly expressed on HSPCs and downregulated during HSPC differentiation. Our characterization of CD82 function using CD82-blocking antibodies revealed a significant decrease in adhesion of HSPCs to niche cells as well as in the in vivo homing and engraftment capabilities of these cells. To determine the molecular mechanisms of CD82s role in adhesion, we have generated CD82 overexpression and knockdown cell lines using the KG1a background. Our data indicate that the level of CD82 expression positively correlates with the extent of adhesion to fibronectin and osteoblasts but has no effect on binding to collagen I or laminin. The increase in adhesion we observed with CD82 overexpression was inhibited by the VLA-4-specific peptide, LDV, indicating a potential role for the VLA-4 (α4β1) integrin. Investigations into potential CD82-mediated mechanisms of VLA-4 regulation have revealed that CD82 regulates both the expression and avidity of VLA-4 but does not regulate its affinity. Taken together, the VLA-4 expression and avidity changes could account for the observed adhesion changes with differing CD82 expression levels. Finally, assessment of CD82 palmitoylation using KG1a CD82 palmitoylation mutant cells revealed that palmitoylation may be required for the CD82-induced changes in adhesion

Attachment styles and psychological profiles of child sex offenders in Ireland

When 29 child sex offenders, 30 violent offenders, 30 nonviolent offenders, and 30 community controls were compared, a secure adult attachment style was 4 times less common in the child sex offender group than in any of the other three groups. Ninety-three percent of sex offenders had an insecure adult attachment style. Compared with community controls, the child sex offender group reported significantly lower levels of maternal and paternal care and significantly higher levels of maternal and paternal overprotection during their childhood. Compared with all three comparison groups, the child sexual offenders reported significantly more emotional loneliness and a more external locus of control. With respect to anger management, the child sexual offenders’ profile more closely approximated those of nonviolent offenders and community controls than that of violent offenders.Author has checked copyright4/12/2013. SB

The lipidated connexin mimetic peptide SRPTEKT-Hdc is a potent inhibitor of Cx43 channels with specificity for the pS368 phospho-isoform

Connexin (Cx) mimetic peptides derived from extracellular loop II sequences (e.g., Gap27: SRPTEKTIFII; Peptide5: VDCFLSRPTEKT) have been used as reversible, Cx-specific blockers of hemichannel (HCh) and gap junction channel (GJCh) function. These blockers typically require high concentrations (~5 µM, 1 h for GJCh) to achieve inhibition. We have shown that addition of a hexadecyl (Hdc) lipid tail to the conserved SRPTEKT peptide sequence (SRPTEKT-Hdc) results in a novel, highly efficacious, and potent inhibitor of mechanically induced Ca2+-wave propagation (IC50 64.8 pM) and HCh-mediated dye uptake (IC50 45.0 pM) in Madin-Darby canine kidney cells expressing rat Cx43 (MDCK43). The lack of similar effect on dye coupling (NBD-MTMA) suggested channel conformation-specific inhibition. Here we report that SRPTEKT-Hdc inhibition of Ca2+-wave propagation, dye coupling, and dye uptake depended on the functional configuration of Cx43 as determined by phosphorylation at serine 368 (S368). Ca2+-wave propagation was enhanced in MDCK cells expressing single-site mutants of Cx43 that mimicked (MDCK43-S368D) or favored (MDCK43-S365A) phosphorylation at S368. Furthermore, SRPTEKT-Hdc potently inhibited GJCh-mediated Ca2+-wave propagation (IC50 230.4 pM), dye coupling, and HCh-mediated dye uptake in MDCK43-S368D and -S365A cells. In contrast, Ca2+-wave propagation, dye coupling, and dye uptake were largely unaffected (IC50 12.3 μM) by SRPTEKT-Hdc in MDCK43-S368A and -S365D cells, mutations that mimic or favor dephosphorylation at S368. Together, these data indicate that SRPTEKT-Hdc is a potent inhibitor of physiological Ca2+-wave signaling mediated specifically by the pS368 phosphorylated form of Cx43.United States Department of Health & Human Services National Institutes of Health (NIH) - USA [HL-058732, HL-131712, HL-007249, NS-073664, GM-055632]12 month embargo; published online: 7 October 2019This item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]

Pancreatic Cancer 3D Cell Line Organoids (CLOs) Maintain the Phenotypic Characteristics of Organoids and Accurately Reflect the Cellular Architecture and Heterogeneity In Vivo

Pancreatic cancer is a highly lethal disease. Therapeutic resistance to chemotherapy is a major cause of treatment failure and recurrence in pancreatic cancer. Organoids derived from cancer stem cells (CSC) are promising models for the advancement of personalised therapeutic responses to inform clinical decisions. However, scaling-up of 3D organoids for high-throughput screening is time-consuming and costly. Here, we successfully developed organoid-derived cell lines (2.5D) from 3D organoids; the cells were then expanded and recapitulated back into organoids known as cell line organoids (CLOs). The 2.5D lines were cultured long term into 2D established cell lines for downstream comparison analysis. Experimental characterisation of the models revealed that the proliferation of CLOs was slightly faster than that of parental organoids. The therapeutic response to chemotherapeutic agents in 3D CLOs and organoids showed a similar responsive profile. Compared to 3D CLOs and organoids, 2D cell lines tended to be less responsive to all the drugs tested. Stem cell marker expression was higher in either 3D CLOs or organoids compared to 2D cell lines. An in vivo tumorigenicity study found CLOs form tumours at a similar rate to organoids and retain enhanced CSC marker expression, indicating the plasticity of CSCs within the in vivo microenvironment