_:b436264121 . _:b436264162 . _:b436264120 . . _:b436264123 . _:b436264122 . _:b436264158 . _:b436264133 . _:b436264132 . . _:b436264135 . . _:b436264123 . . _:b436264134 . _:b436264170 . . . _:b436264150 . _:b436264129 . . _:b8036195 "Cells were prepared for flow cytometry, and at least 50,000 events were acquired on BD Biosciences LSRII or FACSCalibur instruments (BD Biosciences) (14,>>15<<,20). For rare populations, we acquired at least 500 specific events. Data analysis was performed with FlowJo (Tree Star, Ashland, OR) software." . . _:b436264128 . _:b436264177 . _:b436264175 . _:b436264131 . _:b8036195 . _:b436264132 . _:b436264122 . _:b436264130 . . _:b436264141 . . . . _:b436264140 . _:b436264120 . _:b436264143 . _:b436264182 . _:b436264142 . _:b8036191 "The phenotypes of murine cells were determined using four color analyses (>>15<<). Anti-human FoxP3 (eBioscience, San Diego, CA), anti-human CD123 and BDCA2 (Miltenyi Biotec, Bergisch Gladbach, Germany), anti-human CD11c (Biolegend, San Diego, CA), and anti-human CD235a (Coulter Immunotech, Miami, FL) were obtained as" . _:b436264137 . _:b436264118 . . _:b436264136 . _:b436264139 . . . _:b436264138 . _:b436264147 . . _:b436264149 . . . . _:b436264181 . _:b436264148 . _:b436264151 . . . . _:b436264150 . . _:b436264145 . _:b436264144 . . _:b436264147 . _:b436264146 . . _:b436264116 . _:b436264167 . _:b436264157 . . _:b436264139 . _:b436264156 . _:b436264160 . _:b436264159 . . _:b436264158 . _:b436264145 . _:b436264153 . _:b436264125 . _:b436264152 . _:b436264173 . _:b436264155 . _:b436264111 "3"^^ . _:b8036184 . _:b436264152 . _:b436264154 . _:b436264153 . _:b436264168 . _:b436264110 "4"^^ . _:b8036189 . . _:b436264165 . _:b436264164 . _:b8036179 . _:b436264167 . . _:b436264166 . _:b436264157 . . _:b8036189 "research design and methods" . . _:b436264144 . _:b436264161 . _:b436264117 "2"^^ . _:b436264130 . _:b8036187 . . . _:b436264116 "2"^^ . _:b436264160 . _:b8036187 "In separate studies, HSC-engrafted NSG mice readily reject human skin allografts (M.A.B., unpublished observations), but it is known that skin allografts prompt a more robust alloimmune response than do islet allografts (>>24<<,25). Alternatively, the human immune system may be impaired as a result of the hyperglycemic environment\u2014or we may not have allowed sufficient time for graft rejection." . _:b436264119 "2"^^ . _:b436264163 . . _:b436264154 . . _:b436264118 "2"^^ . _:b436264162 . . _:b436264113 "2"^^ . _:b8036197 "Handpicked BALB/c islets (21) were also dissociated into single-cell suspensions (>>22<<). Dissociated islet cell suspensions were treated with Cytofix/Cytoperm (BD Biosciences) for intracellular staining using a biotinylated anti-insulin antibody and developed with Streptavidin-allophycocyanin (SA-APC)." . _:b436264173 . . . _:b436264172 . _:b436264112 "3"^^ . . . _:b436264115 "2"^^ . _:b436264175 . _:b8036189 _:b8036190 . . _:b8036189 _:b8036191 . _:b436264114 "2"^^ . _:b436264174 . _:b436264125 "2"^^ . _:b436264169 . _:b8036184 _:b8036188 . _:b436264168 . _:b436264124 "2"^^ . _:b8036197 . _:b436264127 "2"^^ . _:b436264171 . _:b8036184 _:b8036185 . _:b8036184 _:b8036186 . _:b8036184 _:b8036187 . _:b8036193 "Bone marrow, spleen, and thymus cell suspensions and heparinized blood were incubated with anti-mouse FcR11b to block Fc binding (14,>>19<<). Cells were prepared for flow cytometry, and at least 50,000 events were acquired on BD Biosciences LSRII or FACSCalibur instruments (BD Biosciences) (14,15,20)." . _:b436264126 "2"^^ . _:b436264170 . _:b436264141 . _:b436264121 "2"^^ . _:b436264181 . _:b436264149 . _:b436264180 . _:b436264120 "2"^^ . _:b436264111 . _:b436264174 . _:b436264123 "2"^^ . _:b436264183 . _:b8036189 _:b8036192 . _:b436264182 . _:b436264122 "2"^^ . _:b8036189 _:b8036193 . _:b8036189 _:b8036194 . _:b8036189 _:b8036195 . _:b8036189 _:b8036196 . _:b436264177 . . _:b436264133 "2"^^ . _:b8036189 _:b8036197 . _:b8036189 _:b8036198 . _:b8036189 _:b8036199 . . _:b436264132 "2"^^ . _:b436264176 . . _:b8036179 _:b8036180 . _:b436264179 . _:b436264135 "2"^^ . _:b8036179 _:b8036181 . _:b8036179 _:b8036182 . _:b8036179 _:b8036183 . _:b436264134 "2"^^ . _:b436264178 . . _:b436264129 "2"^^ . _:b436264128 "2"^^ . _:b436264131 "2"^^ . _:b436264130 "2"^^ . _:b436264156 . _:b436264165 . _:b436264141 "2"^^ . _:b8036181 . _:b436264185 . . _:b436264140 "2"^^ . _:b436264184 . _:b8036199 "Pancreata and islet transplants were stained with hemotoxlin-eosin (H-E) and for insulin, glucagon, and human CD45 (>>16<<). Unilateral nephrectomy of the graft-bearing kidney was performed on selected cohorts to confirm recurrent hyperglycemia." . _:b436264143 "2"^^ . _:b436264142 "2"^^ . _:b8036179 "results" . _:b436264137 "2"^^ . _:b436264131 . . _:b8036190 . _:b436264136 "2"^^ . "10.2337%2Fdb10-0323" . . _:b436264139 "2"^^ . _:b8036191 . _:b436264142 . _:b436264169 . _:b436264138 "2"^^ . . _:b436264114 . _:b436264149 "2"^^ . _:b436264148 "2"^^ . _:b436264151 "2"^^ . . . _:b436264150 "2"^^ . . . _:b436264152 . _:b436264153 . _:b436264154 . _:b436264145 "2"^^ . . _:b436264155 . _:b8036180 . _:b436264156 . _:b436264157 . _:b436264164 . _:b436264158 . _:b436264133 . _:b436264144 "2"^^ . _:b436264159 . _:b8036196 . _:b8036181 . _:b436264144 . _:b436264145 . . _:b436264146 . _:b436264147 "2"^^ . . _:b8036182 . _:b436264147 . _:b436264148 . _:b436264149 . _:b8036181 "Although cells with an NK phenotype (DX5+/CD122+) were observed, these cells are not mature NK cells (>>15<<). Populations of granulocytes and macrophages in NRG and NRG-Akita mice were similar (supplemental Table 1), documenting that NRG-Akita mice are deficient in adaptive immunity and NK cells." . _:b436264150 . _:b436264146 "2"^^ . _:b436264151 . _:b8036183 . _:b436264136 . . _:b8036196 "Handpicked BALB/c islets (>>21<<) were also dissociated into single-cell suspensions (22)." . _:b436264127 . _:b436264137 . _:b436264138 . _:b436264157 "2"^^ . _:b436264139 . _:b436264140 . _:b436264141 . _:b436264142 . _:b436264156 "2"^^ . _:b436264143 . _:b436264128 . _:b436264129 . _:b436264130 . _:b436264159 "2"^^ . _:b436264131 . _:b436264132 . _:b436264133 . _:b436264134 . _:b436264158 "2"^^ . _:b436264135 . _:b8036179 . _:b436264184 . _:b436264137 . _:b436264185 . _:b8036190 "Mice were housed in a specific pathogen-free facility in microisolator cages (>>15<<). All animal use was in accordance with the guidelines of the IACUCs of the University of Massachusetts and The Jackson Laboratory." . _:b436264153 "2"^^ . _:b8036188 . _:b436264152 "2"^^ . _:b8036189 . _:b436264176 . _:b436264177 . _:b436264178 . _:b436264155 "2"^^ . _:b436264179 . _:b8036190 . _:b436264180 . _:b436264181 . _:b436264182 . _:b8036182 "We have shown that NRG mice engraft with human HSC at levels similar to those observed in NSG mice (>>14<<), but the effect of chronic hyperglycemia on the development of a human immune system is unknown." . _:b436264159 . _:b436264154 "2"^^ . _:b436264183 . _:b8036191 . _:b8036183 "NRG-Akita and NRG littermates were injected with human HSCs (>>14<<,20). Twelve weeks later, hyperglycemia was confirmed in NRG-Akita mice (data not shown)." . _:b436264168 . _:b436264169 . _:b436264170 . _:b436264165 "2"^^ . _:b436264171 . _:b8036184 . _:b436264172 . _:b436264173 . _:b436264151 . _:b8036183 . _:b436264164 "2"^^ . _:b436264174 . _:b436264175 . _:b8036185 . _:b436264160 . _:b436264161 . _:b436264184 . _:b436264162 . _:b8036182 . _:b436264167 "2"^^ . _:b436264163 . _:b8036186 . _:b436264110 . _:b436264164 . _:b436264165 . _:b436264183 . _:b436264166 . _:b436264166 "2"^^ . _:b436264167 . _:b8036187 . _:b436264161 "2"^^ . _:b8036196 . _:b436264138 . _:b436264160 "2"^^ . _:b8036197 . . _:b436264163 "2"^^ . . _:b8036198 . _:b436264162 "2"^^ . _:b8036199 . . _:b436264173 "2"^^ . . _:b8036192 . . . _:b436264172 "2"^^ . _:b436264179 . _:b8036193 . _:b436264115 . _:b8036194 "Cells were prepared for flow cytometry, and at least 50,000 events were acquired on BD Biosciences LSRII or FACSCalibur instruments (BD Biosciences) (>>14<<,15,20). For rare populations, we acquired at least 500 specific events. Data analysis was performed with FlowJo (Tree Star, Ashland, OR) software." . _:b436264175 "2"^^ . . _:b8036194 . . _:b436264174 "2"^^ . . _:b8036195 . _:b436264120 . _:b436264121 . _:b436264122 . _:b436264169 "2"^^ . _:b436264123 . _:b436264134 . _:b436264124 . _:b436264143 . _:b436264125 . _:b436264126 . _:b436264168 "2"^^ . _:b436264127 . _:b436264112 . _:b436264113 . _:b436264114 . _:b436264171 "2"^^ . _:b436264115 . . _:b436264116 . _:b436264113 . _:b436264117 . _:b436264118 . _:b436264170 "2"^^ . _:b436264119 . _:b436264148 . _:b8036186 . _:b436264181 "2"^^ . . . . . _:b436264110 . _:b436264180 "2"^^ . . _:b436264111 . . _:b436264183 "2"^^ . _:b8036198 . _:b8036186 "We have previously described the development of spontaneously diabetic NOD-Rag1null Prf1null Ins2Akita mice (10), but NOD-Rag1null Prf1null mice engraft poorly with human immune systems (>>23<<). To address this limitation, we generated NRG-Akita mice that spontaneously develop hyperglycemia." . _:b436264185 . _:b8036198 "HSCs were provided by the University of Massachusetts Memorial Umbilical Cord Blood Donation Program under insitutional review board approval (>>14<<,20). Mice were analyzed phenotypically 12\u201316 weeks later and/or transplanted with human islet allografts." . . _:b436264182 "2"^^ . _:b436264135 . _:b436264171 . _:b436264177 "2"^^ . _:b436264161 . . _:b436264176 "2"^^ . _:b8036188 "In separate studies, HSC-engrafted NSG mice readily reject human skin allografts (M.A.B., unpublished observations), but it is known that skin allografts prompt a more robust alloimmune response than do islet allografts (24,>>25<<). Alternatively, the human immune system may be impaired as a result of the hyperglycemic environment\u2014or we may not have allowed sufficient time for graft rejection." . _:b436264179 "2"^^ . _:b8036194 . . _:b8036188 . _:b436264178 "2"^^ . . _:b436264121 . _:b8036192 . _:b436264117 . _:b436264146 . . . _:b436264185 "2"^^ . . _:b436264184 "2"^^ . . "PMC0" . _:b436264180 . _:b436264129 . . . _:b436264124 . _:b8036193 . . . . _:b8036192 "Bone marrow, spleen, and thymus cell suspensions and heparinized blood were incubated with anti-mouse FcR11b to block Fc binding (>>14<<,19). Cells were prepared for flow cytometry, and at least 50,000 events were acquired on BD Biosciences LSRII or FACSCalibur instruments (BD Biosciences) (14,15,20)." . _:b436264172 . _:b436264111 . _:b436264178 . _:b436264163 . _:b8036180 . _:b436264110 . _:b8036184 "discussion" . _:b436264176 . . . _:b436264128 . _:b436264112 . _:b436264117 . _:b436264116 . . _:b436264119 . . _:b436264118 . _:b436264113 . _:b436264136 . _:b8036185 "We have previously described the development of spontaneously diabetic NOD-Rag1null Prf1null Ins2Akita mice (>>10<<), but NOD-Rag1null Prf1null mice engraft poorly with human immune systems (23). To address this limitation, we generated NRG-Akita mice that spontaneously develop hyperglycemia." . _:b436264126 . _:b436264112 . _:b436264155 . . _:b8036199 . _:b436264115 . _:b436264114 . _:b436264140 . _:b8036185 . _:b436264125 . _:b436264124 . _:b436264119 . . _:b8036180 "Because mice homozygous for Ins2Akita develop a rapid hyperglycemia and die shortly after weaning (>>10<<), all studies were performed using mice heterozygous for the Ins2Akita allele." . _:b436264127 . . _:b436264126 . _:b436264166 .