Transgenic Expression of a Novel Secreted and Active Form of IL-33 Promotes Tissue Eosinophilia in a Mouse Model of Eosinophilic Esophagitis

Abstract Although the prevalence of eosinophilic gastrointestinal disorders (EGIDs) is increasing, there is evidence that eosinophilic gastritis and/or duodenitis (EG/EoD) are underdiagnosed. Patients with EG/EoD often present with chronic, non-specific gastrointestinal (GI) symptoms, similar to patients with functional GI disorders. We hypothesized that systematic evaluation, including multiple esophageal, gastric and duodenal biopsies, of patients with chronic GI symptoms might reveal a high rate of gastroduodenal eosinophila, with or without eosinophilic esophagitis (EoE). Methods We performed a prospective multi-center study of patients with non-specific GI symptoms for ≥6 months, from 20 sites. Patients completed a questionnaire assessing abdominal pain, abdominal cramping, early satiety, bloating, nausea, vomiting, diarrhea, and loss of appetite. Those with daily average symptom severity scores ≥3/10 for any single symptom underwent esophagogastroduodenoscopy (EGD) with collection of 4 esophageal (EoE), 8 gastric, and 4 duodenal biopsies, analyzed by central pathologists. Histologic criteria for EoE was ≥15eos/hpf in ≥1 esophageal site and for EG/EoD was peak eosinophil counts ≥30/hpf in ≥5 gastric hpfs and/or 3 duodenal hpfs—criteria used in randomized trials. Results Of 556 patients screened, 405 (73%) met symptom criteria and underwent EGD; 181 patients (45%, mean age 45, 73% female) who underwent EGD met the histologic criteria for EG/EoD, and of these, 7% also had EoE diagnosed. Overall 2% met histologic criteria for EoE alone. Of patients who met the histologic criteria for EG/EoD, 93% were previously diagnosed with gastroesophageal reflux disease (GERD), irritable bowel syndrome (IBS), or functional dyspepsia (FD) (Figure 1). The average duration of GI symptoms in the screened population as well as those that met histologic criteria for EG/EoD was 11 years. Conclusion Forty-five percent of patients with moderate-to-severe GI symptoms who underwent EGD met histologic criteria for EG/EoD. Over 90% of these patients had previously been diagnosed with GERD, IBS, and/or FD, and had minimal overlap with EoE. EGD with systematic gastroduodenal biopsies, and intentional evaluation for tissue eosinophilia, should be performed in patients with chronic GI symptoms. Accurate diagnosis of EG/EoD is required for appropriate, targeted treatment and improved outcomes of patients with moderate-to-severe GI symptoms.

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Siglec-F Inhibition Reduces Esophageal Eosinophilia and Angiogenesis in a Mouse Model of Eosinophilic Esophagitis

Journal of Pediatric Gastroenterology and Nutrition ◽

10.1097/mpg.0b013e3182182ff8 ◽

2011 ◽

Vol 53 (4) ◽

pp. 409-416 ◽

Cited By ~ 57

Author(s):

Eitan Rubinstein ◽

Jae Youn Cho ◽

Peter Rosenthal ◽

James Chao ◽

Marina Miller ◽

...

Keyword(s):

Mouse Model ◽

Eosinophilic Esophagitis ◽

Esophageal Eosinophilia

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Transgenic Expression of Matrix Metalloproteinase-1 Inhibits Myocardial Fibrosis and Prevents the Transition to Heart Failure in a Pressure Overload Mouse Model

Hypertension Research ◽

10.1291/hypres.31.725 ◽

2008 ◽

Vol 31 (4) ◽

pp. 725-735 ◽

Cited By ~ 34

Author(s):

Robert F Foronjy ◽

Jie Sun ◽

Vincent Lemaitre ◽

Jeanine M D'armiento

Keyword(s):

Heart Failure ◽

Mouse Model ◽

Matrix Metalloproteinase ◽

Myocardial Fibrosis ◽

Pressure Overload ◽

Transgenic Expression ◽

Matrix Metalloproteinase 1

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Molecular mechanism of lysosomal sialidase deficiency in galactosialidosis involves its rapid degradation

Biochemical Journal ◽

10.1042/bj3300641 ◽

1998 ◽

Vol 330 (2) ◽

pp. 641-650 ◽

Cited By ~ 38

Author(s):

V. Maia VINOGRADOVA ◽

Lorraine MICHAUD ◽

V. Alexander MEZENTSEV ◽

E. Kiven LUKONG ◽

Mohamed EL-ALFY ◽

...

Keyword(s):

Molecular Mechanism ◽

Half Life ◽

Plasma Membranes ◽

Active Form ◽

Lysosomal Storage ◽

Transgenic Expression ◽

Cultured Skin ◽

Amino Acid Signal Peptide ◽

Amino Acid Signal ◽

Lysosomal Sialidase

Galactosialidosis is an inherited lysosomal storage disease caused by the combined deficiency of lysosomal sialidase and β-galactosidase secondary to the deficiency of cathepsin A/protective protein, which is associated with sialidase and β-galactosidase in a high-molecular weight (1.27 MDa) complex. Clinical phenotypes of patients as well as the composition of compounds which are stored in patient's tissues implicate sialidase deficiency as the underlying pathogenic defect. The recent cloning and sequencing of lysosomal sialidase [Pshezhetsky, Richard, Michaud, Igdoura, Wang, Elsliger, Qu, Leclerc, Gravel, Dallaire and Potier (1997), Nature Genet. 15, 316-320] allowed us to study the molecular mechanism of sialidase deficiency in galactosialidosis. By Western blotting, using antibodies against the recombinant human enzyme, and by NH2-terminal sequencing, we showed that sialidase is synthesized as a 45.5 kDa precursor and after the cleavage of the 47-amino acid signal peptide and glycosylation becomes a 48.3 kDa mature active enzyme present in the 1.27 kDa complex. Transgenic expression of sialidase in cultured skin fibroblasts from normal controls and from galactosialidosis patients, followed by immunofluorescent and immunoelectron microscopy showed that in both normal and affected cells the expressed sialidase was localized on lysosomal and plasma membranes, but the amount of sialidase found in galactosialidosis cells was ~ 5-fold reduced. Metabolic labelling studies demonstrated that the 48.3 kDa mature active form of sialidase was stable in normal fibroblasts (half-life ~ 2.7 h), whereas in galactosialidosis fibroblasts the enzyme was rapidly converted (half-life ~ 30 min) into 38.7 and 24 kDa catalytically inactive forms. Altogether our data provide evidence that the molecular mechanism of sialidase deficiency in galactosialidosis is associated with abnormal proteolytic cleavage and fast degradation.

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Transgenic Expression of Cacna1f Rescues Vision and Retinal Morphology in a Mouse Model of Congenital Stationary Night Blindness 2A (CSNB2A)

Translational Vision Science & Technology ◽

10.1167/tvst.9.11.19 ◽

2020 ◽

Vol 9 (11) ◽

pp. 19

Author(s):

Derek M. Waldner ◽

Kenichi Ito ◽

Li-Li Chen ◽

Lisa Nguyen ◽

Robert L. Chow ◽

...

Keyword(s):

Mouse Model ◽

Night Blindness ◽

Congenital Stationary Night Blindness ◽

Transgenic Expression

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Investigating the role of the ghrelin axis in cognitive decline and dementia

10.23889/suthesis.57625 ◽

2021 ◽

Author(s):

◽

Martina Sassi

Keyword(s):

Mouse Model ◽

Cognitive Decline ◽

Young Animal ◽

Active Form ◽

Hippocampal Neurogenesis ◽

Adult Hippocampal Neurogenesis ◽

Tg2576 Mouse ◽

Brain Functions ◽

Acyl Ghrelin ◽

The Brain

Ghrelin is a 28-amino acid hormone that is generated in a wide number of tissues. Its active form, acyl-ghrelin is able to bind to its receptor GHS-R and exert a variety of functions. In the brain, acyl-ghrelin has been associated with neuroprotection, improved memory and adult hippocampal neurogenesis (AHN). However, the mechanisms controlling acyl-ghrelin-mediated AHN are still unknown. To elucidate this process, different markers of neurogenesis were assessed in a mouse model in which GHS-R+ neurones were ablated specifically from the rostral DG of the hippocampus (rDG), showing that rDG GHS-R+ neurones are essential for maintaining AHN. Acyl-ghrelin has also been shown to prevents the damage caused by neurodegeneration, at least in young animal models of disease. In this thesis, we demonstrated that in the geriatric Tg2576 mouse model of Alzheimer’s disease, acyl-ghrelin maintained an effect on β-amyloid (Aβ) plaques in the hippocampus, promoting a reduction of the Aβ plaques size in AD-like mouse model compared to WT mice. Collectively, research findings highlight the importance of circulating acyl-ghrelin in the brain. However, ghrelin exists in two distinct forms and acyl-ghrelin can be enzymatically modified to the ‘inactive' unacylated-ghrelin (UAG) by acyl-protein thioesterase 1 (APT1). Preventing APT1 mediated de-acylation and increasing acyl-ghrelin bio-availability may prevent the damage caused by neurodegeneration. Unpublished data from our group suggest that PalmostatinB, an APT1 inhibitor, increases levels of acyl-ghrelin in macrophage cells (that naturally produce ghrelin). Therefore, this and other APT1 inhibitors may be considered possible therapeutic agents for the treatment of cognitive decline and diseases associated with dementia. We confirmed that, among several APT1 inhibitors, PalmostatinB is able to increase the level of acyl-ghrelin in vitro. However, further research is warranted into APT1 inhibitors as a novel therapeutic approach to treating cognitive decline and dementia. Together, the data in this thesis support a role for the ghrelinergic system components in modulating brain functions.

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Transgenic Expression of Inducible Caspase9 Suicide Gene for In Vivo Elimination of Antigen Specific Cytotoxic T Cells (CTLs) Engineered To Produce Cytokines.

Blood ◽

10.1182/blood.v108.11.137.137 ◽

2006 ◽

Vol 108 (11) ◽

pp. 137-137

Author(s):

C. Quintarelli ◽

V. Juan ◽

B. Savoldo ◽

G. Giordano ◽

A. Foster ◽

...

Keyword(s):

T Cells ◽

Mouse Model ◽

Scid Mouse ◽

Cytotoxic T Cells ◽

Safety Margin ◽

Suicide Gene ◽

Cytokine Release ◽

Transgenic Expression ◽

Scid Mouse Model

Abstract Transgenic expression of cytokines such as IL2 and IL15 by tumor specific cytotoxic T cells (CTLs) can improve the survival and expansion of these cells within the tumor microenvironment. Using our model of tumor antigen-Epstein-Barr virus-specific CTLs (EBV-CTLs), we found that transgenic production of either cytokine could sustain CTL expansion ex vivo, and improve their anti-tumor effect in vivo in a SCID mouse model. Although the proliferation of cytokine gene transduced CTLs remained antigen dependent, clinical application of this approach likely requires the inclusion of a suicide gene to deal with the potential development of T-cell mutants with autonomous, antigen-independent, growth. We constructed a novel suicide gene based on the inducible caspase9 gene [Straathof et al Blood 2005:105;4247] and showed that EBV-CTLs expressing this suicide gene can be eliminated after exposure to the small molecule chemical inducer of dimerization (CID) AP20187. However, it has been suggested that the anti-apoptotic effects of transgenic cytokine expression would counteract the pro-apoptotic action of icaspase9. To discover the general applicability of our inducible suicide gene approach, we generated tricistronic retroviral vectors encoding full length hIL2 or hIL15, a truncated human CD34 molecule (DCD34) as a marker, and the inducible Caspase9 suicide gene. The three genes were linked using 2A peptide cleavable sequences, which allows equal expression of the 3 transgenes. Established EBV-CTLs were transduced either with IL2.DCD34.iCasp9 or IL15.DCD34.iCasp9 or DCD34 empty vector. After stimulation with the antigen (LCLs) at a E:T ratio of 1:1, we observed significant expansion of CTLs/IL2.DCD34.iCasp9 (221 fold expansion, range 84–452) and IL15.DCD34.iCasp9 (182, range 48–355) after 28 days of culture, while CTLs/DCD34 maintained without cytokines had <2 fold expansion. Cytokine release after antigen stimulation was 6.4±3.4 pg/mL and 157±99pg/mL for IL15 and IL2, respectively. Transgenic CTLs maintained the same phenotype as control T cells (>90% CD3+/CD8+) and remained polyclonal as assessed by staining for the TCRVb repertoire. To evaluate the cytotoxic capacity of the suicide gene, control and transgenic CTLs were incubated with CID AP20187 at 20nM. After 24 hours the survival of transgenic CTLs was estimated from residual CD34+ expression. For both CTLs/IL2.DCD34.iCasp9 and CTLs/IL15.DCD34.iCasp9 the percentage of CD34+ cells was reduced by >1.5 logs. Moreover, the few residual cells were CD34dim, and produced no measurable cytokine release as measured by ELISA. To assess activity in vivo, we used an LCL-engrafted SCID mouse model, injected with CTLs transgenic for the firefly-luciferase gene. These cells were tracked in vivo using the Xenogen-IVIS bioluminescence system. We found that CTLs transgenic for either IL2 or IL15 both migrated to the tumor site, and had increased expansion compared to DCD34 control CTLs. Mice were then treated with 2 −3 doses of CID AP20187 (50 mg/mice i.p). The bioluminescence signal diminished to background levels by 48hr, suggesting efficient elimination of transgenic CTLs. In conclusion, these data indicate that induction of a transgenic caspase9 suicide gene can effectively destroy adoptively transferred T cells, even when these lymphocytes are expressing anti-apoptotic cytokines such as IL2 and IL15. This approach may increase the safety margin of gene modified CTL therapies.

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The NLRP3 inflammasome inhibitor OLT1177 rescues cognitive impairment in a mouse model of Alzheimer’s disease

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2009680117 ◽

2020 ◽

Vol 117 (50) ◽

pp. 32145-32154

Author(s):

Niklas Lonnemann ◽

Shirin Hosseini ◽

Carlo Marchetti ◽

Damaris B. Skouras ◽

Davide Stefanoni ◽

...

Keyword(s):

Alzheimer’S Disease ◽

Alzheimer's Disease ◽

Synaptic Plasticity ◽

Mouse Model ◽

Nlrp3 Inflammasome ◽

Therapeutic Potential ◽

Active Form ◽

Specific Inhibitor ◽

Morris Water Maze Test ◽

Model Of Alzheimer’S Disease

Numerous studies demonstrate that neuroinflammation is a key player in the progression of Alzheimer’s disease (AD). Interleukin (IL)-1β is a main inducer of inflammation and therefore a prime target for therapeutic options. The inactive IL-1β precursor requires processing by the the nucleotide-binding oligomerization domain-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome into a mature and active form. Studies have shown that IL-1β is up-regulated in brains of patients with AD, and that genetic inactivation of the NLRP3 inflammasome improves behavioral tests and synaptic plasticity phenotypes in a murine model of the disease. In the present study, we analyzed the effect of pharmacological inhibition of the NLRP3 inflammasome using dapansutrile (OLT1177), an oral NLRP3-specific inhibitor that is safe in humans. Six-month-old WT and APP/PS1 mice were fed with standard mouse chow or OLT1177-enriched chow for 3 mo. The Morris water maze test revealed an impaired learning and memory ability of 9-mo-old APP/PS1 mice (P = 0.001), which was completely rescued by OLT1177 fed to mice (P = 0.008 to untreated APP/PS1). Furthermore, our findings revealed that 3 mo of OLT1177 diet can rescue synaptic plasticity in this mouse model of AD (P = 0.007 to untreated APP/PS1). In addition, microglia were less activated (P = 0.07) and the number of plaques was reduced in the cortex (P = 0.03) following NLRP3 inhibition with OLT1177 administration. We also observed an OLT1177 dose-dependent normalization of plasma metabolic markers of AD to those of WT mice. This study suggests the therapeutic potential of treating neuroinflammation with an oral inhibitor of the NLRP3 inflammasome.

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