Tonya Bliss - Academia.edu (original) (raw)

Papers by Tonya Bliss

Stroke, Feb 1, 2013

Introduction: The mechanisms of functional recovery after stroke are thought to be based on struc... more Introduction: The mechanisms of functional recovery after stroke are thought to be based on structural and functional changes in brain circuits adjacent to or connected with the stroke site. Deciphering these changes at the synaptic level is key to understanding the re-organization of the synaptic circuitry. Here we use a combined approach of i) array tomography to determine the composition of GABA synapses in the post-stroke mouse brain, with ii) electrophysiology to determine whether stroke leads to functional changes in GABA A receptor-mediated neurotransmission. Methods: A cortical lesion was induced in 12-week-old C57BL/6J male mice using the distal middle cerebral artery occlusion model of ischemia. For array tomography, small tissue was removed from the peri-infarct cortex and ribbons of serial ultrathin sections were obtained. Ribbons were stained with antibodies for synaptic markers. Analysis of the resultant staining pattern was used to quantify GABAergic synapses. In addition, whole-cell patch clamp recordings from acute neocortical brain slices were performed to evaluate GABA-mediated synaptic signaling in the peri-infarct cortex. Behavior was evaluated weekly. Results: At 1 week post-stroke, the array tomography data revealed an increase in the density and proportion of alpha1 subunit-containing GABAergic synapses in layer 5 of the peri-infarct cortex (Density: 0.064 vs 0.036 synapses/μm3. Proportion: 15.3 vs 9.1 %, p<0.05, n=6); no changes were observed in layer2/3. Changes in GABA synapses were transient and returned to basal levels by 1 month. Electrophysiological recordings at 1 week post-stroke showed that GABA A receptor-mediated currents were enhanced in layer 5, but not in layer 2/3. These changes were specific to the pyramidal neurons. Behavioral impairment after stroke was observed only at 1 week compared to sham mice (p<0.05, n=10). Conclusion: Our results suggest that stroke leads to an increased expression of functional GABA A receptors in peri-infarct neocortex and that these changes are layer- and cell type-specific. These synaptic changes may represent a mechanism of post-stroke functional recovery and remapping of surviving circuits.

Cell Reports, Apr 1, 2023

Stroke, 2013

Introduction: The mechanisms of functional recovery after stroke are thought to be based on struc... more Introduction: The mechanisms of functional recovery after stroke are thought to be based on structural and functional changes in brain circuits adjacent to or connected with the stroke site. Deciphering these changes at the synaptic level is key to understanding the re-organization of the synaptic circuitry. Here we use a combined approach of i) array tomography to determine the composition of GABA synapses in the post-stroke mouse brain, with ii) electrophysiology to determine whether stroke leads to functional changes in GABA A receptor-mediated neurotransmission. Methods: A cortical lesion was induced in 12-week-old C57BL/6J male mice using the distal middle cerebral artery occlusion model of ischemia. For array tomography, small tissue was removed from the peri-infarct cortex and ribbons of serial ultrathin sections were obtained. Ribbons were stained with antibodies for synaptic markers. Analysis of the resultant staining pattern was used to quantify GABAergic synapses. In addi...

Stroke, 2013

Inflammation critically contributes to post-stroke brain damage. Mast cells (MCs), perivascular c... more Inflammation critically contributes to post-stroke brain damage. Mast cells (MCs), perivascular cells best known as effector cells involved in the development of inflammatory processes, have been reported to exacerbate stroke pathology. Unlike other immune cells, mature MCs do not circulate but are resident in virtually all anatomical sites, including brain parenchyma and meninges. Thus a key question is which tissue-specific MCs are important after stroke. To address this, we used ‘mast cell knock-in’ mouse models whereby genetically MC-deficient mice were selectively repaired of their MC deficiency by engraftment of in vitro grown mast cells. Methods: Two different MC-deficient mouse models (KitW-sh/W-sh and KitW/W-v) were used. For each model, 3 groups were tested: wild-type, MC-deficient, and MC-engrafted. Mice were subjected to 30 min occlusion of the middle cerebral artery. Brain swelling and infarct size were assessed by T2-weighted MRI and histology. The immune response was ...

Functional recovery after stroke is thought to involve structural plasticity in circuits adjacent... more Functional recovery after stroke is thought to involve structural plasticity in circuits adjacent to the lesion (1). Deciphering these changes at the synaptic level is key to understanding the re-organization of the synaptic circuitry. Here we describe the use of array tomography (2, 3), a new highresolution imaging method, to determine the composition of glutamate and GABA synapses in the post-stroke mouse brain. Electrophysiology was done to determine if these quantitative changes resulted in functional changes.

Stroke, 2022

Background: Acute hyperglycemia occurs in over 40% of ischemic stroke patients, increases hemorrh... more Background: Acute hyperglycemia occurs in over 40% of ischemic stroke patients, increases hemorrhagic transformation (HT) and worsens stroke outcome. Previous rodent studies reported deleterious effects of hyperglycemia on stroke outcome during acute stroke (within hours to days), however, its impact during subacute stroke period (days to weeks) remains unclear. Moreover, the mechanisms underlying hyperglycemia’s worsening of stroke outcomes remains elusive. In this study, we investigated the effect of acute hyperglycemia on immune responses and stroke outcome in acute and subacute phases. Method: Male C57/BL6 mice were subjected to middle cerebral artery occlusion (MCAO) for 30 min, followed by reperfusion to mimic ischemic stroke. Acute hyperglycemia was induced by glucose injection 10 min before MCAO. For the acute phase study, mice were sacrificed at 4.5 hrs to assess blood-brain barrier leakage (Evans blue) and brain immune cell populations (flow cytometry), and at 24 hrs to ev...

Stroke, 2016

Introduction: Neural stem cell (NSC) transplantation is being explored as a potential therapy for... more Introduction: Neural stem cell (NSC) transplantation is being explored as a potential therapy for ischemic stroke (IS). Although research has shown the efficacy of NSCs in IS, little is known about...

Functional recovery after stroke is thought to involve structural plasticity in circuits adjacent... more Functional recovery after stroke is thought to involve structural plasticity in circuits adjacent to the lesion (1). Deciphering these changes at the synaptic level is key to understanding the re-organization of the synaptic circuitry. Here we describe the use of array tomography (2, 3), a new highresolution imaging method, to determine the composition of glutamate and GABA synapses in the post-stroke mouse brain. Electrophysiology was done to determine if these quantitative changes resulted in functional changes.

Cell transplantation offers a novel therapeutic strategy for stroke; however, how transplanted ce... more Cell transplantation offers a novel therapeutic strategy for stroke; however, how transplanted cells function in vivo is poorly understood. We show for the first time that after sub-acute transplantation into the ischemic brain of human central nervous system stem cells grown as neurospheres (hCNS-SCns), the stem cell-secreted factor, human VEGF (hVEGF), is necessary for cell-induced functional recovery. We correlate this functional recovery to hVEGF-induced effects on the host brain including multiple facets of vascular repair, and its unexpected suppression of the inflammatory response. We found that transplanted hCNS-SCns affected multiple parameters in the brain with different kinetics: early improvement in blood-brain barrier (BBB) integrity and suppression of inflammation was followed by a delayed spatio-temporal regulated increase in neovascularization. These events coincided with a bi-modal pattern of functional recovery: an early recovery independent of neovascularization, ...

Stroke, 2013

Introduction: Stem cell transplantation has emerged as a promising new experimental treatment for... more Introduction: Stem cell transplantation has emerged as a promising new experimental treatment for stroke; understanding its mechanism of action will facilitate the translation of stem cell therapy to the clinic. Previous work from our lab and others suggests that transplanted stem cells function by enhancing endogenous brain repair processes including structural brain plasticity. The ultimate change in brain plasticity is manifested at the synaptic level and thus we hypothesize that stem cells will enhance synaptic structural remodeling in the post-ischemic brain. To test this we use array tomography, a new high-resolution proteomic imaging method, to determine a) the number and subtype of glutamate and GABA synapses after stroke, and b) how these parameters are affected by transplantation of human neural progenitor cells (hNPCs). Method: Vehicle or hNPCs derived from fetal cortex were transplanted into the ischemic cortex of Nude rats at 7 days after distal middle cerebral artery o...

We have previously characterized human neuronal progenitor cells (hNP) that can adopt a retinal g... more We have previously characterized human neuronal progenitor cells (hNP) that can adopt a retinal ganglion cell (RGC)-like morphology within the RGC and nerve fiber layers of the retina. In an effort to determine whether hNPs could be used a candidate cells for targeted delivery of neurotrophic factors (NTFs), we evaluated whether hNPs transfected with an vector that expresses IGF-1 in the form of a fusion protein with tdTomato (TD), would increase RGC survival in vitro and confer neuroprotective effects in a mouse model of glaucoma. RGCs co-cultured with hNP IGF-TD cells displayed enhanced survival, and increased neurite extension and branching as compared to hNP TD or untransfected hNP cells. Application of various IGF-1 signaling blockers or IGF-1 receptor antagonists abrogated these effects. In vivo, using a model of glaucoma we showed that IOP elevation led to reductions in retinal RGC count. In this model, evaluation of retinal flatmounts and optic nerve cross sections indicated that only hNP IGF-TD cells effectively reduced RGC death and showed a trend to improve optic nerve axonal loss. RT-PCR analysis of retina lysates over time showed that the neurotrophic effects of IGF-1 were also attributed to down-regulation of inflammatory and to some extent, angiogenic pathways. This study shows that neuronal progenitor cells that hone into the RGC and nerve fiber layers may be used as vehicles for local production and delivery of a desired NTF. Transplantation of hNP IGF-TD cells improves RGC survival in vitro and protects against RGC loss in a rodent model of glaucoma. Our findings have provided experimental evidence and form the basis for applying cell-based strategies for local delivery of NTFs into the retina. Application of cell-based delivery may be extended to other disease conditions beyond glaucoma.

Stroke, 2014

Background: Stroke is a major cause of disability yet pharmacotherapy targeting the recovery phas... more Background: Stroke is a major cause of disability yet pharmacotherapy targeting the recovery phase is lacking. Cortical circuit reorganization adjacent to the stroke site promotes recovery, thus elucidating mechanisms that promote this plasticity could lead to new therapeutics. Tonic neuronal inhibition, mediated by extrasynaptic GABAA receptors,inhibits post-stroke recovery. However, effects of phasic (synaptic) GABA signaling - which promotes plasticity during development - are unknown. Here we use a combined approach of i) array tomography to determine the composition of GABA synapses in the post-stroke mouse brain, ii) electrophysiology to determine whether stroke leads to functional changes in GABA-mediated phasic inhibition, and (iii) treatment with zolpidem, an FDA-approved GABA agonist, to modulate recovery. Results: We found, using array tomography, a 1.7-fold increase in the number of GABAergic synapses containing the α1 receptor subunit in layer 5 of the peri-infarct cort...

Stroke, 2012

Introduction: The mechanisms of functional recovery after stroke are thought to be based on struc... more Introduction: The mechanisms of functional recovery after stroke are thought to be based on structural and functional changes in brain circuits adjacent to or connected with the stroke site. Deciphering these changes at the synaptic level is key to understanding the re-organization of the synaptic circuitry (i.e. the connectome). Quantitative information about such synapse rearrangements after stroke has been inadequate however, due to the technical limitations of available methodologies. Here we describe the use of array tomography, a new high-resolution proteomic imaging method, to determine the composition of glutamate and GABA synapses in the post-stroke mouse brain. Methods: A cortical lesion was induced in 12-week-old C57BL/6J male mice using the distal middle cerebral artery occlusion model of ischemia. Small tissue sections were removed from the peri-infarct cortex and ribbons of serial ultrathin (70 nm) sections were obtained using an ultramicrotome. Ribbons were stained wi...

Stroke, 2014

Background: Inflammation critically contributes to post-stroke brain damage and consequently ther... more Background: Inflammation critically contributes to post-stroke brain damage and consequently there is much interest in factors that influence stroke-induced brain inflammation. Interleukin 6 (IL-6) has been heralded as an important predictor of stroke outcome, however there are conflicting reports describing detrimental, beneficial and even no effects of IL-6 on stroke pathology. This has led to the suggestion that the post-stroke effects of IL-6 may be dependent on its cellular location. We report that IL-6 produced by meningeal resident mast cells (pro-inflammatory effector cells) significantly exacerbates brain inflammation and pathology after stroke. These are the first data to identify the role in stroke pathology of IL-6 derived from a specific cell type in a particular anatomic location in vivo. Methods: A mast cell (MC) ‘knock-in’ mouse model was used where MC-deficient mice (KitW/W-v) were repaired of their MC deficiency by engraftment of in-vitro-derived MCs. MCs from wild...

Stroke, Feb 1, 2013

Introduction: The mechanisms of functional recovery after stroke are thought to be based on struc... more Introduction: The mechanisms of functional recovery after stroke are thought to be based on structural and functional changes in brain circuits adjacent to or connected with the stroke site. Deciphering these changes at the synaptic level is key to understanding the re-organization of the synaptic circuitry. Here we use a combined approach of i) array tomography to determine the composition of GABA synapses in the post-stroke mouse brain, with ii) electrophysiology to determine whether stroke leads to functional changes in GABA A receptor-mediated neurotransmission. Methods: A cortical lesion was induced in 12-week-old C57BL/6J male mice using the distal middle cerebral artery occlusion model of ischemia. For array tomography, small tissue was removed from the peri-infarct cortex and ribbons of serial ultrathin sections were obtained. Ribbons were stained with antibodies for synaptic markers. Analysis of the resultant staining pattern was used to quantify GABAergic synapses. In addition, whole-cell patch clamp recordings from acute neocortical brain slices were performed to evaluate GABA-mediated synaptic signaling in the peri-infarct cortex. Behavior was evaluated weekly. Results: At 1 week post-stroke, the array tomography data revealed an increase in the density and proportion of alpha1 subunit-containing GABAergic synapses in layer 5 of the peri-infarct cortex (Density: 0.064 vs 0.036 synapses/μm3. Proportion: 15.3 vs 9.1 %, p<0.05, n=6); no changes were observed in layer2/3. Changes in GABA synapses were transient and returned to basal levels by 1 month. Electrophysiological recordings at 1 week post-stroke showed that GABA A receptor-mediated currents were enhanced in layer 5, but not in layer 2/3. These changes were specific to the pyramidal neurons. Behavioral impairment after stroke was observed only at 1 week compared to sham mice (p<0.05, n=10). Conclusion: Our results suggest that stroke leads to an increased expression of functional GABA A receptors in peri-infarct neocortex and that these changes are layer- and cell type-specific. These synaptic changes may represent a mechanism of post-stroke functional recovery and remapping of surviving circuits.

Cell Reports, Apr 1, 2023

Stroke, 2013

Introduction: The mechanisms of functional recovery after stroke are thought to be based on struc... more Introduction: The mechanisms of functional recovery after stroke are thought to be based on structural and functional changes in brain circuits adjacent to or connected with the stroke site. Deciphering these changes at the synaptic level is key to understanding the re-organization of the synaptic circuitry. Here we use a combined approach of i) array tomography to determine the composition of GABA synapses in the post-stroke mouse brain, with ii) electrophysiology to determine whether stroke leads to functional changes in GABA A receptor-mediated neurotransmission. Methods: A cortical lesion was induced in 12-week-old C57BL/6J male mice using the distal middle cerebral artery occlusion model of ischemia. For array tomography, small tissue was removed from the peri-infarct cortex and ribbons of serial ultrathin sections were obtained. Ribbons were stained with antibodies for synaptic markers. Analysis of the resultant staining pattern was used to quantify GABAergic synapses. In addi...

Stroke, 2013

Inflammation critically contributes to post-stroke brain damage. Mast cells (MCs), perivascular c... more Inflammation critically contributes to post-stroke brain damage. Mast cells (MCs), perivascular cells best known as effector cells involved in the development of inflammatory processes, have been reported to exacerbate stroke pathology. Unlike other immune cells, mature MCs do not circulate but are resident in virtually all anatomical sites, including brain parenchyma and meninges. Thus a key question is which tissue-specific MCs are important after stroke. To address this, we used ‘mast cell knock-in’ mouse models whereby genetically MC-deficient mice were selectively repaired of their MC deficiency by engraftment of in vitro grown mast cells. Methods: Two different MC-deficient mouse models (KitW-sh/W-sh and KitW/W-v) were used. For each model, 3 groups were tested: wild-type, MC-deficient, and MC-engrafted. Mice were subjected to 30 min occlusion of the middle cerebral artery. Brain swelling and infarct size were assessed by T2-weighted MRI and histology. The immune response was ...

Functional recovery after stroke is thought to involve structural plasticity in circuits adjacent... more Functional recovery after stroke is thought to involve structural plasticity in circuits adjacent to the lesion (1). Deciphering these changes at the synaptic level is key to understanding the re-organization of the synaptic circuitry. Here we describe the use of array tomography (2, 3), a new highresolution imaging method, to determine the composition of glutamate and GABA synapses in the post-stroke mouse brain. Electrophysiology was done to determine if these quantitative changes resulted in functional changes.

Stroke, 2022

Background: Acute hyperglycemia occurs in over 40% of ischemic stroke patients, increases hemorrh... more Background: Acute hyperglycemia occurs in over 40% of ischemic stroke patients, increases hemorrhagic transformation (HT) and worsens stroke outcome. Previous rodent studies reported deleterious effects of hyperglycemia on stroke outcome during acute stroke (within hours to days), however, its impact during subacute stroke period (days to weeks) remains unclear. Moreover, the mechanisms underlying hyperglycemia’s worsening of stroke outcomes remains elusive. In this study, we investigated the effect of acute hyperglycemia on immune responses and stroke outcome in acute and subacute phases. Method: Male C57/BL6 mice were subjected to middle cerebral artery occlusion (MCAO) for 30 min, followed by reperfusion to mimic ischemic stroke. Acute hyperglycemia was induced by glucose injection 10 min before MCAO. For the acute phase study, mice were sacrificed at 4.5 hrs to assess blood-brain barrier leakage (Evans blue) and brain immune cell populations (flow cytometry), and at 24 hrs to ev...

Stroke, 2016

Introduction: Neural stem cell (NSC) transplantation is being explored as a potential therapy for... more Introduction: Neural stem cell (NSC) transplantation is being explored as a potential therapy for ischemic stroke (IS). Although research has shown the efficacy of NSCs in IS, little is known about...

Functional recovery after stroke is thought to involve structural plasticity in circuits adjacent... more Functional recovery after stroke is thought to involve structural plasticity in circuits adjacent to the lesion (1). Deciphering these changes at the synaptic level is key to understanding the re-organization of the synaptic circuitry. Here we describe the use of array tomography (2, 3), a new highresolution imaging method, to determine the composition of glutamate and GABA synapses in the post-stroke mouse brain. Electrophysiology was done to determine if these quantitative changes resulted in functional changes.

Cell transplantation offers a novel therapeutic strategy for stroke; however, how transplanted ce... more Cell transplantation offers a novel therapeutic strategy for stroke; however, how transplanted cells function in vivo is poorly understood. We show for the first time that after sub-acute transplantation into the ischemic brain of human central nervous system stem cells grown as neurospheres (hCNS-SCns), the stem cell-secreted factor, human VEGF (hVEGF), is necessary for cell-induced functional recovery. We correlate this functional recovery to hVEGF-induced effects on the host brain including multiple facets of vascular repair, and its unexpected suppression of the inflammatory response. We found that transplanted hCNS-SCns affected multiple parameters in the brain with different kinetics: early improvement in blood-brain barrier (BBB) integrity and suppression of inflammation was followed by a delayed spatio-temporal regulated increase in neovascularization. These events coincided with a bi-modal pattern of functional recovery: an early recovery independent of neovascularization, ...

Stroke, 2013

Introduction: Stem cell transplantation has emerged as a promising new experimental treatment for... more Introduction: Stem cell transplantation has emerged as a promising new experimental treatment for stroke; understanding its mechanism of action will facilitate the translation of stem cell therapy to the clinic. Previous work from our lab and others suggests that transplanted stem cells function by enhancing endogenous brain repair processes including structural brain plasticity. The ultimate change in brain plasticity is manifested at the synaptic level and thus we hypothesize that stem cells will enhance synaptic structural remodeling in the post-ischemic brain. To test this we use array tomography, a new high-resolution proteomic imaging method, to determine a) the number and subtype of glutamate and GABA synapses after stroke, and b) how these parameters are affected by transplantation of human neural progenitor cells (hNPCs). Method: Vehicle or hNPCs derived from fetal cortex were transplanted into the ischemic cortex of Nude rats at 7 days after distal middle cerebral artery o...

We have previously characterized human neuronal progenitor cells (hNP) that can adopt a retinal g... more We have previously characterized human neuronal progenitor cells (hNP) that can adopt a retinal ganglion cell (RGC)-like morphology within the RGC and nerve fiber layers of the retina. In an effort to determine whether hNPs could be used a candidate cells for targeted delivery of neurotrophic factors (NTFs), we evaluated whether hNPs transfected with an vector that expresses IGF-1 in the form of a fusion protein with tdTomato (TD), would increase RGC survival in vitro and confer neuroprotective effects in a mouse model of glaucoma. RGCs co-cultured with hNP IGF-TD cells displayed enhanced survival, and increased neurite extension and branching as compared to hNP TD or untransfected hNP cells. Application of various IGF-1 signaling blockers or IGF-1 receptor antagonists abrogated these effects. In vivo, using a model of glaucoma we showed that IOP elevation led to reductions in retinal RGC count. In this model, evaluation of retinal flatmounts and optic nerve cross sections indicated that only hNP IGF-TD cells effectively reduced RGC death and showed a trend to improve optic nerve axonal loss. RT-PCR analysis of retina lysates over time showed that the neurotrophic effects of IGF-1 were also attributed to down-regulation of inflammatory and to some extent, angiogenic pathways. This study shows that neuronal progenitor cells that hone into the RGC and nerve fiber layers may be used as vehicles for local production and delivery of a desired NTF. Transplantation of hNP IGF-TD cells improves RGC survival in vitro and protects against RGC loss in a rodent model of glaucoma. Our findings have provided experimental evidence and form the basis for applying cell-based strategies for local delivery of NTFs into the retina. Application of cell-based delivery may be extended to other disease conditions beyond glaucoma.

Stroke, 2014

Background: Stroke is a major cause of disability yet pharmacotherapy targeting the recovery phas... more Background: Stroke is a major cause of disability yet pharmacotherapy targeting the recovery phase is lacking. Cortical circuit reorganization adjacent to the stroke site promotes recovery, thus elucidating mechanisms that promote this plasticity could lead to new therapeutics. Tonic neuronal inhibition, mediated by extrasynaptic GABAA receptors,inhibits post-stroke recovery. However, effects of phasic (synaptic) GABA signaling - which promotes plasticity during development - are unknown. Here we use a combined approach of i) array tomography to determine the composition of GABA synapses in the post-stroke mouse brain, ii) electrophysiology to determine whether stroke leads to functional changes in GABA-mediated phasic inhibition, and (iii) treatment with zolpidem, an FDA-approved GABA agonist, to modulate recovery. Results: We found, using array tomography, a 1.7-fold increase in the number of GABAergic synapses containing the α1 receptor subunit in layer 5 of the peri-infarct cort...

Stroke, 2012

Introduction: The mechanisms of functional recovery after stroke are thought to be based on struc... more Introduction: The mechanisms of functional recovery after stroke are thought to be based on structural and functional changes in brain circuits adjacent to or connected with the stroke site. Deciphering these changes at the synaptic level is key to understanding the re-organization of the synaptic circuitry (i.e. the connectome). Quantitative information about such synapse rearrangements after stroke has been inadequate however, due to the technical limitations of available methodologies. Here we describe the use of array tomography, a new high-resolution proteomic imaging method, to determine the composition of glutamate and GABA synapses in the post-stroke mouse brain. Methods: A cortical lesion was induced in 12-week-old C57BL/6J male mice using the distal middle cerebral artery occlusion model of ischemia. Small tissue sections were removed from the peri-infarct cortex and ribbons of serial ultrathin (70 nm) sections were obtained using an ultramicrotome. Ribbons were stained wi...

Stroke, 2014

Background: Inflammation critically contributes to post-stroke brain damage and consequently ther... more Background: Inflammation critically contributes to post-stroke brain damage and consequently there is much interest in factors that influence stroke-induced brain inflammation. Interleukin 6 (IL-6) has been heralded as an important predictor of stroke outcome, however there are conflicting reports describing detrimental, beneficial and even no effects of IL-6 on stroke pathology. This has led to the suggestion that the post-stroke effects of IL-6 may be dependent on its cellular location. We report that IL-6 produced by meningeal resident mast cells (pro-inflammatory effector cells) significantly exacerbates brain inflammation and pathology after stroke. These are the first data to identify the role in stroke pathology of IL-6 derived from a specific cell type in a particular anatomic location in vivo. Methods: A mast cell (MC) ‘knock-in’ mouse model was used where MC-deficient mice (KitW/W-v) were repaired of their MC deficiency by engraftment of in-vitro-derived MCs. MCs from wild...