A flexible loop in the paxillin LIM3 domain mediates its direct binding to integrin ß subunits.

Integrins are fundamental for cell adhesion and the formation of focal adhesions (FA). Accordingly, these receptors guide embryonic development, tissue maintenance, and haemostasis but are also involved in cancer invasion and metastasis. A detailed understanding of the molecular interactions that drive integrin activation, FA assembly, and downstream signalling cascades is critical. Here, we reveal a direct association of paxillin, a marker protein of FA sites, with the cytoplasmic tails of the integrin ß1 and ß3 subunits. The binding interface resides in paxillin's LIM3 domain, where based on the NMR structure and functional analyses, a flexible, 7-amino acid loop engages the unstructured part of the integrin cytoplasmic tail. Genetic manipulation of the involved residues in either paxillin or integrin ß3 compromises cell adhesion and motility of murine fibroblasts. This direct interaction between paxillin and the integrin cytoplasmic domain identifies an alternative, kindlin-independent mode of integrin outside-in signalling particularly important for integrin ß3 function.

SLC44A2-mediated phenotypic switch of vascular smooth muscle cells contributes to aortic aneurysm.

The phenotypic switch of vascular smooth cells (VSMCs) from a contractile to a synthetic state is associated with the development and progression of aortic aneurysm (AA). However, the mechanism underlying this process remains unclear. In this issue of the JCI, Song et al. identified SLC44A2 as a regulator of the phenotypic switch in VSMCs. Inhibition of SLC44A2 facilitated the switch to the synthetic state, contributing to the development of AA. Mechanistically, SLC44A2 interacted with NRP1 and ITGB3 to activate the TGF-ß/SMAD signaling pathway, resulting in VSMCs with a contractile phenotype. Furthermore, VSMC-specific SLC44A2 overexpression by genetic or pharmacological manipulation reduced AA in mouse models. These findings suggest the potential of targeting the SLC44A2 signaling pathway for AA prevention and treatment.

Targeting ITGß3 to Overcome Trastuzumab Resistance through Epithelial-Mesenchymal Transition Regulation in HER2-Positive Breast Cancer.

HER2-positive breast cancer, representing 15-20% of all breast cancer cases, often develops resistance to the HER2-targeted therapy trastuzumab. Unfortunately, effective treatments for advanced HER2-positive breast cancer remain scarce. This study aims to investigate the roles of ITGß3, and Hedgehog signaling in trastuzumab resistance and explore the potential of combining trastuzumab with cilengitide as a therapeutic strategy. Quantitative gene expression analysis was performed to assess the transcription of EMT (epithelial-mesenchymal transition) markers Slug, Snail, Twist2, and Zeb1 in trastuzumab-resistant HER2-positive breast cancer cells. The effects of ITGß3 and Hedgehog signaling were investigated. Additionally, the combination therapy of trastuzumab and cilengitide was evaluated. Acquired trastuzumab resistance induced the transcription of Slug, Snail, Twist2, and Zeb1, indicating increased EMT. This increased EMT was mediated by ITGB3 and Hedgehog signaling. ITGß3 regulated both the Hedgehog pathway and EMT, with the latter being independent of the Hedgehog pathway. The combination of trastuzumab and cilengitide showed a synergistic effect, reducing both EMT and Hedgehog pathway activity. Targeting ITGß3 with cilengitide, combined with trastuzumab, effectively suppresses the Hedgehog pathway and EMT, offering a potential strategy to overcome trastuzumab resistance and improve outcomes for HER2-positive breast cancer patients.

Integrin ß3 enhances glycolysis and increases lactate production in endometriosis.

BACKGROUND: Endometriosis (EMs) is a chronic disease characterized by endometrial-like tissue present outside of the uterus. Macrophages have been confirmed to participate in the development of EMs. Integrin ß3 (ITGB3), a ß-subunit of the integrin family, is crucial in tumor progression. In this study, we investigated the pivotal role of ITGB3 in endometrial stromal cells (ESCs) and its influence on the development of EMs, particularly focusing on the regulatory impact of macrophages. METHODS: In this study, we used western blot, Real-time qPCR, Immunohistochemistry to detected the high expression of ITGB3 in ESCs. ITGB3-overexpression ESCs (ITGB3-OE) was constructed and detected by RNA-seq with normal ESCs. ATP and lactate expression assay, transwell migration assay, wound healing, cell adhesion assay and other molecular biology techniques were used to explore the potential mechanisms. In vivo, we constructed the EMs mouse model and injected with cilengitite to inhibit ITGB3. RESULTS: Here, we found ITGB3 highly expressed in ectopic lesions in EMs. The increasing ITGB3 resulted in activating the glycolysis, which produced more ATP and lactate in ITGB3-OE. After culturing with lactate, the migration, proliferation and invasion ability of ESCs were enhanced, while the result in 2-DG was reversed. In vivo, the results showed that after antagonizing ITGB3, the number of ectopic lesions was decrease. CONCLUSIONS: Our findings indicate that ITGB3 up-regulated by macrophages are able to regulate the glycolysis to promote the development of EMs and lactate enhances the ability of proliferation, migration, invasion and adhesion of EMs iv vivo and in vitro.

FGF1 Suppresses Allosteric Activation of ß3 Integrins by FGF2: A Potential Mechanism of Anti-Inflammatory and Anti-Thrombotic Action of FGF1.

Several inflammatory cytokines bind to the allosteric site (site 2) and allosterically activate integrins. Site 2 is also a binding site for 25-hydroxycholesterol, an inflammatory lipid mediator, and is involved in inflammatory signaling (e.g., TNF and IL-6 secretion) in addition to integrin activation. FGF2 is pro-inflammatory and pro-thrombotic, and FGF1, homologous to FGF2, has anti-inflammatory and anti-thrombotic actions, but the mechanism of these actions is unknown. We hypothesized that FGF2 and FGF1 bind to site 2 of integrins and regulate inflammatory signaling. Here, we describe that FGF2 is bound to site 2 and allosterically activated ß3 integrins, suggesting that the pro-inflammatory action of FGF2 is mediated by binding to site 2. In contrast, FGF1 bound to site 2 but did not activate these integrins and instead suppressed integrin activation induced by FGF2, indicating that FGF1 acts as an antagonist of site 2 and that the anti-inflammatory action of FGF1 is mediated by blocking site 2. A non-mitogenic FGF1 mutant (R50E), which is defective in binding to site 1 of αvß3, suppressed ß3 integrin activation by FGF2 as effectively as WT FGF1.

Overcoming Vemurafenib Resistance in Metastatic Melanoma: Targeting Integrins to Improve Treatment Efficacy.

Metastatic melanoma, a deadly form of skin cancer, often develops resistance to the BRAF inhibitor drug vemurafenib, highlighting the need for understanding the underlying mechanisms of resistance and exploring potential therapeutic strategies targeting integrins and TGF-ß signalling. In this study, the role of integrins and TGF-ß signalling in vemurafenib resistance in melanoma was investigated, and the potential of combining vemurafenib with cilengitide as a therapeutic strategy was investigated. In this study, it was found that the transcription of PAI1 and p21 was induced by acquired vemurafenib resistance, and ITGA5 levels were increased as a result of this resistance. The transcription of ITGA5 was mediated by the TGF-ß pathway in the development of vemurafenib resistance. A synergistic effect on the proliferation of vemurafenib-resistant melanoma cells was observed with the combination therapy of vemurafenib and cilengitide. Additionally, this combination therapy significantly decreased invasion and colony formation in these resistant cells. In conclusion, it is suggested that targeting integrins and TGF-ß signalling, specifically ITGA5, ITGB3, PAI1, and p21, may offer promising approaches to overcoming vemurafenib resistance, thereby improving outcomes for metastatic melanoma patients.

PEX3 promotes regenerative repair after myocardial injury in mice through facilitating plasma membrane localization of ITGB3.

The peroxisome is a versatile organelle that performs diverse metabolic functions. PEX3, a critical regulator of the peroxisome, participates in various biological processes associated with the peroxisome. Whether PEX3 is involved in peroxisome-related redox homeostasis and myocardial regenerative repair remains elusive. We investigate that cardiomyocyte-specific PEX3 knockout (Pex3-KO) results in an imbalance of redox homeostasis and disrupts the endogenous proliferation/development at different times and spatial locations. Using Pex3-KO mice and myocardium-targeted intervention approaches, the effects of PEX3 on myocardial regenerative repair during both physiological and pathological stages are explored. Mechanistically, lipid metabolomics reveals that PEX3 promotes myocardial regenerative repair by affecting plasmalogen metabolism. Further, we find that PEX3-regulated plasmalogen activates the AKT/GSK3ß signaling pathway via the plasma membrane localization of ITGB3. Our study indicates that PEX3 may represent a novel therapeutic target for myocardial regenerative repair following injury.

Role of TSP-1 and its receptor ITGB3 in the renal tubulointerstitial injury of focal segmental glomerulosclerosis.

Focal segmental glomerulosclerosis (FSGS), a common cause of primary glomerulonephritis, has a poor prognosis and is pathologically featured by tubulointerstitial injury. Thrombospondin-1 (TSP-1) is an extracellular matrix protein that acts in combination with different receptors in the kidney. Here, we analyzed the tubular expression of TSP-1 and its receptor integrin ß3 (ITGB3) in FSGS. Previously the renal interstitial chip analysis of FSGS patients with tubular interstitial injury showed that the expression of TSP-1 and ITGB3 were upregulated. We found that the expression of TSP-1 and ITGB3 increased in the tubular cells of FSGS patients. The plasma level of TSP-1 increased and was correlated to the degree of tubulointerstitial lesions in FSGS patients. TSP-1/ITGB3 signaling induced renal tubular injury in HK-2 cells exposure to bovine serum albumin and the adriamycin (ADR)-induced nephropathy model. THBS1 KO ameliorated tubular injury and renal fibrosis in ADR-treated mice. THBS1 knockdown decreased the expression of KIM-1 and caspase 3 in the HK-2 cells treated with bovine serum albumin, while THBS1 overexpression could induce tubular injury. In vivo, we identified cyclo-RGDfK as an agent to block the binding of TSP-1 to ITGB3. Cyclo-RGDfK treatment could alleviate ADR-induced renal tubular injury and interstitial fibrosis in mice. Moreover, TSP-1 and ITGB3 were colocalized in tubular cells of FSGS patients and ADR-treated mice. Taken together, our data showed that TSP-1/ITGB3 signaling contributed to the development of renal tubulointerstitial injury in FSGS, potentially identifying a new therapeutic target for FSGS.

The Role of TLR-2 in Lethal COVID-19 Disease Involving Medullary and Resident Lung Megakaryocyte Up-Regulation in the Microthrombosis Mechanism.

Patients with COVID-19 have coagulation and platelet disorders, with platelet alterations and thrombocytopenia representing negative prognostic parameters associated with severe forms of the disease and increased lethality. METHODS: The aim of this study was to study the expression of platelet glycoprotein IIIa (CD61), playing a critical role in platelet aggregation, together with TRL-2 as a marker of innate immune activation. RESULTS: A total of 25 patients were investigated, with the majority (24/25, 96%) having co-morbidities and dying from a fatal form of SARS-CoV-2(+) infection (COVID-19+), with 13 men and 12 females ranging in age from 45 to 80 years. When compared to a control group of SARS-CoV-2 (-) negative lungs (COVID-19-), TLR-2 expression was up-regulated in a subset of patients with deadly COVID-19 fatal lung illness. The proportion of Spike-1 (+) patients found by PCR and ISH correlates to the proportion of Spike-S1-positive cases as detected by digital pathology examination. Furthermore, CD61 expression was considerably higher in the lungs of deceased patients. In conclusion, we demonstrate that innate immune prolonged hyperactivation is related to platelet/megakaryocyte over-expression in the lung. CONCLUSIONS: Microthrombosis in deadly COVID-19+ lung disease is associated with an increase in the number of CD61+ platelets and megakaryocytes in the pulmonary interstitium, as well as their functional activation; this phenomenon is associated with increased expression of innate immunity TLR2+ cells, which binds the SARS-CoV-2 E protein, and significantly with the persistence of the Spike-S1 viral sequence.

4'-O-MethylbavachalconeB Targeted 14-3-3ζ Blocking the Integrin ß3 Early Outside-In Signal to Inhibit Platelet Aggregation and Thrombosis.

14-3-3ζ protein, the key target in the regulation and control of integrin ß3 outside-in signaling, is an attractive new strategy to inhibit thrombosis without affecting hemostasis. In this study, 4'-O-methylbavachalconeB (4-O-MB) in Psoraleae Fructus was identified as a 14-3-3ζ ligand with antithrombosis activity by target fishing combined with ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS) analysis. The competitive inhibition analysis showed that 4-O-MB targeted 14-3-3ζ and blocked the 14-3-3ζ/integrin ß3 interaction with inhibition constant (Ki) values of 9.98 ± 0.22 µM. Molecular docking and amino acid mutation experiments confirmed that 4-O-MB specifically bound to 14-3-3ζ through LSY9 and SER28 to regulate the 14-3-3ζ/integrin ß3 interaction. Besides, 4-O-MB affected the integrin ß3 early outside-in signal by inhibiting AKT and c-Src phosphorylation. Meanwhile, 4-O-MB could inhibit ADP-, collagen-, or thrombin-induced platelet aggregation function but had no effect on platelet adhesion to collagen-coated surfaces in vivo. Administration of 4-O-MB could significantly inhibit thrombosis formation without disturbing hemostasis in mice. These findings provide new prospects for the antithrombotic effects of Psoraleae Fructus and the potential application of 4-O-MB as lead compounds in the therapy of thrombosis by targeting 14-3-3ζ.

Endometrial BMP2 Deficiency Impairs ITGB3-Mediated Trophoblast Invasion in Women With Repeated Implantation Failure.

BACKGROUND: Repeated implantation failure (RIF) leads to a waste of high-quality embryos and remains a challenge in assisted reproductive technology. During early human placentation, the invasion of trophoblast cells into the decidua is an essential step for the establishment of maternal-fetal interactions and subsequent successful pregnancy. Bone morphogenetic protein 2 (BMP2) has been reported to regulate endometrial receptivity and promote trophoblast invasion. However, whether there is dysregulation of endometrial BMP2 expression in patients with RIF remains unknown. Additionally, the molecular mechanisms underlying the effects of BMP2 on human trophoblast invasion and early placentation remain to be further elucidated. METHODS: Midluteal phase endometrial samples were biopsied from patients with RIF and from routine control in vitro fertilization followed by quantitative polymerase chain reaction and immunoblotting analyses. Human trophoblast organoids, primary human trophoblast cells, and an immortalized trophoblast cell line (HTR8/SVneo) were used as study models. RESULTS: We found that BMP2 was aberrantly low in midluteal phase endometrial tissues from patients with RIF. Recombinant human BMP2 treatment upregulated integrin ß3 (ITGB3) in a SMAD2/3-SMAD4 signaling-dependent manner in both HTR8/SVneo cells and primary trophoblast cells. siRNA-mediated integrin ß3 downregulation reduced both basal and BMP2-upregulated trophoblast invasion and vascular mimicry in HTR8/SVneo cells. Importantly, shRNA-mediated ITGB3 knockdown significantly decreased the formation ability of human trophoblast organoids. CONCLUSION: Our results demonstrate endometrial BMP2 deficiency in patients with RIF. ITGB3 mediates both basal and BMP2-promoted human trophoblast invasion and is essential for early placentation. These findings broaden our knowledge regarding the regulation of early placentation and provide candidate diagnostic and therapeutic targets for RIF clinical management.

The role of thrombomodulin in modulating ITGB3 expression and its implications for triple-negative breast cancer progression.

Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer (BC) compared to other BC subtypes in clinical settings. Currently, there are no effective therapeutic strategies for TNBC treatment. Therefore, there is an urgent need to identify suitable biomarkers or therapeutic targets for TNBC patients. Thrombomodulin (TM) plays a role in cancer progression and metastasis in many different cancers. However, the role of TM in TNBC is not yet fully understood. First, silenced-TM in MDA-MB-231 cells caused an increase in proliferative and metastatic activity. In contrast, overexpression of TM in Hs578T cells caused a reduction in proliferation, invasion, and migration rate. Using RNA-seq analysis, we found that Integrin beta 3 (ITGB3) expression may be a downstream target of TM. Furthermore, we found an increase in ITGB3 levels in TM-KD cells by QPCR and western blot analysis but a decrease in ITGB3 levels in TM-overexpressing cells. We found phospho-smad2/3 levels were increased in TM-KD cells but decreased in TM-overexpressing cells. This implies that TM negatively regulates ITGB3 levels through the activation of the smad2/3 pathway. Silencing ITGB3 in TM-KD cells caused a decrease in proliferation and migration. Finally, we found that higher ITGB3 levels were correlated with poor overall survival and relapse-free survival in patients with TNBC. Our results indicated a novel regulatory relationship between TM and ITGB3 in TNBC.

Effect of c.1431C > T mutation, a causative mutation of Glanzmann's thrombasthenia, on ITGB3 splicing, gene and protein expression.

BACKGROUND/AIM: Recently, it was reported that the non-synonymous c.1431C > T (p. G477=) mutation of the integrin subunit ß3 (ITGB3) gene is the cause of Glanzmann's thrombasthenia (GT). However, the functional consequences of this mutation on the ITGB3 gene and protein expression remain to be elucidated. Therefore, this study was conducted to cover this scientific shortage. METHODS: Peripheral blood samples were collected from Chinese family members (parents and proband and his sister), and DNA was extracted and sequenced using whole-exome and Sanger sequencing. The effect of c.1431C > T mutation on the splicing of mRNA was verified by the in vitro minigene assay and the three variants that resulted from the mutation were cloned into a phage vector and pEGFP-C1 vector, and ITGB3 gene and protein expression was detected in the transfected 293 T cells using qPCR and Western blotting. RESULTS: Minigene splicing assay showed that c.1431C > T mutation causes three kinds of alternative splicing; (1) a 95 bp deletion in the middle of exon10, (2) a 155 bp deletion (95 bp deletion in the middle of exon10 plus a 60 bp deletion in the right side of exon10), and (3) a 261 bp deletion in the right side of exon10. The in vitro expression assay showed that the c.1431C > T variant did not affect the ITGB3 mRNA levels, but directly led to protein truncation and declined expression. CONCLUSION: Due to its significant impact on protein expression, c.1431C > T mutation in ITGB3 could be considered a pathogenic variant of GT. This could enrich the ITGB3 mutation spectrum and provide a base for the genetic diagnosis of GT.

SINE-VNTR-Alu retrotransposon insertion as a novel mutational event underlying Glanzmann thrombasthenia.

BACKGROUND: Glanzmann thrombasthenia (GT) is an autosomal recessive platelet aggregation disorder caused by mutations in ITGA2B or ITGB3. OBJECTIVES: We aimed to assess the phenotype and investigate the genetic etiology of a GT pedigree. METHODS: A patient with bleeding manifestations and mild mental retardation was enrolled. Complete blood count, coagulation, and platelet aggregation tests were performed. Causal mutations were identified via whole exome and genome sequencing and subsequently confirmed through polymerase chain reaction and Sanger sequencing. The transcription of ITGB3 was characterized using RNA sequencing and reverse transcription polymerase chain reaction. The αⅡb and ß3 biosynthesis was investigated via whole blood flow cytometry and in vitro studies. RESULTS: GT was diagnosed in a patient with defective platelet aggregation. Novel compound heterozygous ITGB3 variants were identified, with a maternal nonsense mutation (c.2222G>A, p.Trp741∗) and a paternal SINE-VNTR-Alu (SVA) retrotransposon insertion. The 5' truncated SVA element was inserted in a sense orientation in intron 11 of ITGB3, resulting in aberrant splicing of ITGB3 and significantly reducing ß3 protein content. Meanwhile, both the expression and transportation of ß3 were damaged by the ITGB3 c.2222G>A. Almost no αⅡb and ß3 expressions were detected on the patient's platelets surface. CONCLUSION: Novel compound heterozygous ITGB3 mutations were identified in the GT pedigree, resulting in defects of αⅡbß3 biosynthesis. This is the first report of SVA retrotransposon insertion in the genetic pathogenesis of GT. Our study highlights the importance of combining multiple high-throughput sequencing technologies for the molecular diagnosis of genetic disorders.

Regulation of dendritic spine length in corticopontine layer V pyramidal neurons by autism risk gene ß3 integrin.

The relationship between autism spectrum disorder (ASD) and dendritic spine abnormalities is well known, but it is unclear whether the deficits relate to specific neuron types and brain regions most relevant to ASD. Recent genetic studies have identified a convergence of ASD risk genes in deep layer pyramidal neurons of the prefrontal cortex. Here, we use retrograde recombinant adeno-associated viruses to label specifically two major layer V pyramidal neuron types of the medial prefrontal cortex: the commissural neurons, which put the two cerebral hemispheres in direct communication, and the corticopontine neurons, which transmit information outside the cortex. We compare the basal dendritic spines on commissural and corticopontine neurons in WT and KO mice for the ASD risk gene Itgb3, which encodes for the cell adhesion molecule ß3 integrin selectively enriched in layer V pyramidal neurons. Regardless of the genotype, corticopontine neurons had a higher ratio of stubby to mushroom spines than commissural neurons. ß3 integrin affected selectively spine length in corticopontine neurons. Ablation of ß3 integrin resulted in corticopontine neurons lacking long (> 2 µm) thin dendritic spines. These findings suggest that a deficiency in ß3 integrin expression compromises specifically immature spines on corticopontine neurons, thereby reducing the cortical territory they can sample. Because corticopontine neurons receive extensive local and long-range excitatory inputs before relaying information outside the cortex, specific alterations in dendritic spines of corticopontine neurons may compromise the computational output of the full cortex, thereby contributing to ASD pathophysiology.

Molecular dynamics simulations corroborate recombinant expression studies carried out on three αIIb ß-propeller mutations reported in Indian Glanzmann thrombasthenia patients.

Mutations in the αIIb ß-propeller domain have long been known to disrupt heterodimerization and intracellular trafficking of αIIbß3 complexes leading to diminished surface expression and/or function, resulting in Glanzmann thrombasthenia. Our previous study on three ß-propeller mutations, namely G128S, S287L, and G357S, showed variable defects in protein transport correlated with the patient's clinical phenotypes. Pulse-chase experiments revealed differences in αIIbß3 complex maturation among the three mutations. Hence, the current study aims to correlate conformational changes caused by each one of them. Evolutionary conservation analysis, stability analysis, and molecular dynamics simulations of the three mutant structures were carried out. Stability analysis revealed that, while G128S and G357S mutations destabilized the ß-propeller structure, S287L retained the stability. Wild-type and mutant ß-propeller structures, when subjected to molecular dynamics simulations, confirmed that G128S and G357S were both destabilizing in nature when compared with the wild-type and S287L based on several parameters studied, like RMSD, RMSF, Rg, FEL, PCA, secondary structure, and hydrogen bonds. In our previous study, we demonstrated that mutant S287L αIIbß3 complexes were more stable than the wild-type αIIbß3 complexes, as evidenced in pulse-chase experiments. These findings corroborate variable intracellular fates of mutant αIIbß3 complexes as a result of these ß-propeller mutations.

Effect of acupuncture on the opening time of implantation window and endometrial receptivity in controlled ovarian hyperstimulation rats during peri-implantation period.

Purpose: To investigate the effect of acupuncture for improving the pregnancy rate of COH rats from the viewpoint of regulating the opening time of the implantation window and endometrial receptivity. Methods: Experimental rats were randomly divided into normal group (N), model group (M) and acupuncture group(A), and samples were collected on Day 4, 5 and 6 after mating. COH rats were treated with acupuncture at SP6, LR3, and ST36 once a day for 7 times. The pinopodes were observed under a scanning electron microscope. Serum estrogen and progesterone levels were measured via ELISA. The protein and mRNA levels of estrogen receptor (ER), progesterone receptor (PR), leukemia inhibitory factor (LIF), integrin ß3, vascular endothelial growth factor (VEGF), and fibroblast growth factor 2 (FGF-2) in the endometrium were evaluated via West-blot, immunohistochemistry, and PCR. Results: Compared with group N, the pregnancy rate of group M was significantly decreased (P<0.05), and the abnormal serum hormone levels and implantation window advancement were observed. Compared with group M, the pregnancy rate of group A was significantly increased (P<0.05), the supraphysiological serum progesterone levels were restored to normalcy (P<0.05), and the advanced implantation window was restored to a certain extent. Further, the abnormal ER, PR, LIF, integrin ß3, VEGF, and FGF-2 expression levels of the endometrium got recovered to varying degrees. Conclusion: Acupuncture may restore the estrogen and progesterone balance in COH rats and the forward shift of the implantation window to a certain extent, improving the endometrial receptivity and finally improving the pregnancy rate of COH rats.

Integrin ß3 regulates apical dendritic morphology of pyramidal neurons throughout hippocampal CA3.

In excitatory hippocampal pyramidal neurons, integrin ß3 is critical for synaptic maturation and plasticity in vitro. Itgb3 is a potential autism susceptibility gene that regulates dendritic morphology in the cerebral cortex in a cell-specific manner. However, it is unknown what role Itgb3 could have in regulating hippocampal pyramidal dendritic morphology in vivo, a key feature that is aberrant in many forms of autism and intellectual disability. We found that Itgb3 mRNA is expressed in the stratum pyramidale of CA3. We examined the apical dendritic morphology of CA3 hippocampal pyramidal neurons in conditional Itgb3 knockouts and controls, utilizing the Thy1-GFP-M line. We fully reconstructed the apical dendrite of each neuron and determined each neuron's precise location along the dorsoventral, proximodistal, and radial axes of the stratum pyramidale. We found a very strong effect for Itgb3 expression on CA3 apical dendritic morphology: neurons from conditional Itgb3 knockouts had longer and thinner apical dendrites than controls, particularly in higher branch orders. We also assessed potential relationships between pairs of topographic or morphological variables, finding that most variable pairs were free from any linear relationships to each other. We also found that some neurons from controls, but not conditional Itgb3 knockouts, had a graded pattern of overall diameter along the dorsoventral and proximodistal axes of the stratum pyramidale of CA3. Taken together, Itgb3 is essential for constructing normal dendritic morphology in pyramidal neurons throughout CA3.