Recent genomic studies challenge the conventional model that each metastasis must arise from a single tumor cell and instead reveal that metastases can be composed of multiple genetically distinct ...clones. These intriguing observations raise the question: How do polyclonal metastases emerge from the primary tumor? In this study, we used multicolor lineage tracing to demonstrate that polyclonal seeding by cell clusters is a frequent mechanism in a common mouse model of breast cancer, accounting for >90% of metastases. We directly observed multicolored tumor cell clusters across major stages of metastasis, including collective invasion, local dissemination, intravascular emboli, circulating tumor cell clusters, and micrometastases. Experimentally aggregating tumor cells into clusters induced a >15-fold increase in colony formation ex vivo and a >100-fold increase in metastasis formation in vivo. Intriguingly, locally disseminated clusters, circulating tumor cell clusters, and lung micrometastases frequently expressed the epithelial cytoskeletal protein, keratin 14 (K14). RNA-seq analysis revealed that K14⁺ cells were enriched for desmosome and hemidesmosome adhesion complex genes, and were depleted for MHC class II genes. Depletion of K14 expression abrogated distant metastases and disrupted expression of multiple metastasis effectors, including Tenascin C (Tnc), Jagged1 (Jag1), and Epiregulin (Ereg). Taken together, our findings reveal K14 as a key regulator of metastasis and establish the concept that K14⁺ epithelial tumor cell clusters disseminate collectively to colonize distant organs.
Monte Carlo simulation studies are among the primary scientific outputs contributed by methodologists, guiding application of various statistical tools in practice. Although methodological ...researchers routinely extend simulation study findings through follow-up work, few studies are ever replicated. Simulation studies are susceptible to factors that can contribute to replicability failures, however. This paper sought to conduct a meta-scientific study by replicating one highly cited simulation study (Curran et al., Psychological Methods, 1, 16-29, 1996) that investigated the robustness of normal theory maximum likelihood (ML)-based chi-square fit statistics under multivariate nonnormality. We further examined the generalizability of the original study findings across different nonnormal data generation algorithms. Our replication results were generally consistent with original findings, but we discerned several differences. Our generalizability results were more mixed. Only two results observed under the original data generation algorithm held completely across other algorithms examined. One of the most striking findings we observed was that results associated with the independent generator (IG) data generation algorithm vastly differed from other procedures examined and suggested that ML was robust to nonnormality for the particular factor model used in the simulation. Findings point to the reality that extant methodological recommendations may not be universally valid in contexts where multiple data generation algorithms exist for a given data characteristic. We recommend that researchers consider multiple approaches to generating a specific data or model characteristic (when more than one is available) to optimize the generalizability of simulation results.
The regeneration of oligodendrocytes is a crucial step in recovery from demyelination, as surviving oligodendrocytes exhibit limited structural plasticity and rarely form additional myelin sheaths. ...New oligodendrocytes arise through the differentiation of platelet‐derived growth factor receptor α (PDGFRα) expressing oligodendrocyte progenitor cells (OPCs) that are widely distributed throughout the CNS. Although there has been detailed investigation of the behavior of these progenitors in white matter, recent studies suggest that disease burden in multiple sclerosis (MS) is more strongly correlated with gray matter atrophy. The timing and efficiency of remyelination in gray matter is distinct from white matter, but the dynamics of OPCs that contribute to these differences have not been defined. Here, we used in vivo genetic fate tracing to determine the behavior of OPCs in gray and white matter regions in response to cuprizone‐induced demyelination. Our studies indicate that the temporal dynamics of OPC differentiation varies significantly between white and gray matter. While OPCs rapidly repopulate the corpus callosum and mature into CC1 expressing mature oligodendrocytes, OPC differentiation in the cingulate cortex and hippocampus occurs much more slowly, resulting in a delay in remyelination relative to the corpus callosum. The protracted maturation of OPCs in gray matter may contribute to greater axonal pathology and disease burden in MS.
Main Points
The timing and efficiency of remyelination in gray matter is distinct from white matter.
Our studies indicate that the temporal dynamics of OPC differentiation varies significantly between white and gray matter.
Multiple sclerosis (MS) is a demyelinating disease of the CNS characterized by inflammation and neurodegeneration. Animal models that enable the study of remyelination in the context of ongoing ...inflammation are greatly needed for the development of novel therapies that target the pathological inhibitory cues inherent to the MS plaque microenvironment. We report the development of an innovative animal model combining cuprizone-mediated demyelination with transfer of myelin-reactive CD4(+) T cells. Characterization of this model reveals both Th1 and Th17 CD4(+) T cells infiltrate the CNS of cuprizone-fed mice, with infiltration of Th17 cells being more efficient. Infiltration correlates with impaired spontaneous remyelination as evidenced by myelin protein expression, immunostaining, and ultrastructural analysis. Electron microscopic analysis further reveals that demyelinated axons are preserved but reduced in caliber. Examination of the immune response contributing to impaired remyelination highlights a role for peripheral monocytes with an M1 phenotype. This study demonstrates the development of a novel animal model that recapitulates elements of the microenvironment of the MS plaque and reveals an important role for T cells and peripheral monocytes in impairing endogenous remyelination in vivo. This model could be useful for testing putative MS therapies designed to enhance remyelination in the setting of active inflammation, and may also facilitate modeling the pathophysiology of denuded axons, which has been a challenge in rodents because they typically remyelinate very quickly.
Pharmacologic targeting of T helper (T H) cell trafficking poses an attractive opportunity for amelioration of autoimmune diseases such as multiple sclerosis (MS). MS risk is associated with vitamin ...D deficiency, and its bioactive form, 1,25-dihydroxyvitamin D3 1,25(OH) ₂D ₃, has been shown to prevent experimental autoimmune encephalomyelitis, a mouse model of MS, via an incompletely understood mechanism. Herein, we systematically examined 1,25(OH) ₂D ₃ effects on T H cells during their migration from the lymph nodes to the CNS. Our data demonstrate that myelin-reactive T H cells are successfully generated in the presence of 1,25(OH) ₂D ₃, secrete proinflammatory cytokines, and do not preferentially differentiate into suppressor T cells. These cells are able to leave the lymph node, enter the peripheral circulation, and migrate to the s.c. immunization sites. However, T H cells from 1,25(OH) ₂D ₃-treated mice are unable to enter the CNS parenchyma but are instead maintained in the periphery. Upon treatment cessation, mice rapidly develop experimental autoimmune encephalomyelitis, demonstrating that 1,25(OH) ₂D ₃ prevents the disease only temporarily likely by halting T H cell migration into the CNS.
The mammary epithelium elaborates through hormonally regulated changes in proliferation, migration and differentiation. Non-muscle myosin II (NMII) functions at the interface between contractility, ...adhesion and signal transduction. It is therefore a plausible regulator of mammary morphogenesis. We tested the genetic requirement for NMIIA and NMIIB in mammary morphogenesis through deletion of the three NMII heavy chain-encoding genes (
,
and
; also known as
,
and
, respectively) that confer specificity to the complex. Surprisingly, mosaic loss, but not ubiquitous loss, of
and
induced high levels of proliferation in 3D culture. This phenotype was observed even when cells were cultured in basal medium, which does not support tissue level growth of wild-type epithelium. Mosaic loss of NMIIA and NMIIB combined with FGF signaling to induce hyperplasia. Mosaic analysis revealed that the cells that were null for both NMIIA and NMIIB, as well as wild-type cells, proliferated, indicating that the regulation of proliferation is both cell autonomous and non-autonomous within epithelial tissues. This phenotype appears to be mediated by cell-cell contact, as co-culture did not induce proliferation. Mosaic loss of NMIIA and NMIIB also induced excess proliferation
Our data therefore reveal a role for NMIIA and NMIIB as negative regulators of proliferation in the mammary epithelium.
Nerve conduction within the mammalian central nervous system is made efficient by oligodendrocyte‐derived myelin. Historically, thyroid hormones have a well described role in regulating ...oligodendrocyte differentiation and myelination during development; however, it remains unclear which thyroid hormone receptors are required to drive these effects. This is a question with clinical relevance since nonspecific thyroid receptor stimulation can produce deleterious side‐effects. Here we report that GC‐1, a thyromimetic with selective thyroid receptor β action and a potentially limited side‐effect profile, promotes in vitro oligodendrogenesis from both rodent and human oligodendrocyte progenitor cells. In addition, we used in vivo genetic fate tracing of oligodendrocyte progenitor cells via PDGFαR‐CreER;Rosa26‐eYFP double‐transgenic mice to examine the effect of GC‐1 on cellular fate and find that treatment with GC‐1 during developmental myelination promotes oligodendrogenesis within the corpus callosum, occipital cortex and optic nerve. GC‐1 was also observed to enhance the expression of the myelin proteins MBP, CNP and MAG within the same regions. These results indicate that a β receptor selective thyromimetic can enhance oligodendrocyte differentiation in vitro and during developmental myelination in vivo and warrants further study as a therapeutic agent for demyelinating models. GLIA 2014;62:1513–1529
Main Points:
Thyroid hormones regulate oligodendrocyte differentiation during development, but it remains unclear which receptors are required to drive this effect.
We report that GC‐1, a thyromimetic with selective receptor β action and a limited side‐effect profile, promotes oligodendrogenesis.
Significance Vitamin D plays an important role in regulating the immune system in health and disease and may be beneficial for patients with multiple sclerosis. It prevents CNS autoimmunity in mice ...by an incompletely understood mechanism. The present study is a systematic evaluation of the vitamin D effects on T lymphocytes at each step of their journey to the CNS. The data demonstrate that vitamin D does not affect generation of pathogenic cells but prevents their presence in the CNS. Unlike current long-acting drugs that impair immune cell trafficking, the effect of vitamin D is quickly reversed after treatment cessation, which could prove advantageous when immune function needs to be reestablished in the setting of infection.
Pharmacologic targeting of T helper (T H ) cell trafficking poses an attractive opportunity for amelioration of autoimmune diseases such as multiple sclerosis (MS). MS risk is associated with vitamin D deficiency, and its bioactive form, 1,25-dihydroxyvitamin D3 1,25(OH) 2 D 3 , has been shown to prevent experimental autoimmune encephalomyelitis, a mouse model of MS, via an incompletely understood mechanism. Herein, we systematically examined 1,25(OH) 2 D 3 effects on T H cells during their migration from the lymph nodes to the CNS. Our data demonstrate that myelin-reactive T H cells are successfully generated in the presence of 1,25(OH) 2 D 3 , secrete proinflammatory cytokines, and do not preferentially differentiate into suppressor T cells. These cells are able to leave the lymph node, enter the peripheral circulation, and migrate to the s.c. immunization sites. However, T H cells from 1,25(OH) 2 D 3 -treated mice are unable to enter the CNS parenchyma but are instead maintained in the periphery. Upon treatment cessation, mice rapidly develop experimental autoimmune encephalomyelitis, demonstrating that 1,25(OH) 2 D 3 prevents the disease only temporarily likely by halting T H cell migration into the CNS.
Pharmacologic targeting of T helper (T^sub H^) cell trafficking poses an attractive opportunity for amelioration of autoimmune diseases such as multiple sclerosis (MS). MS risk is associated with ...vitamin D deficiency, and its bioactive form, 1,25-dihydroxyvitamin D3 1,25(OH)^sub 2^D^sub 3^, has been shown to prevent experimental autoimmune encephalomyelitis, a mouse model of MS, via an incompletely understood mechanism. Herein, we systematically examined 1,25(OH)^sub 2^D^sub 3^ effects on T^sub H^ cells during their migration from the lymph nodes to the CNS. Our data demonstrate that myelin-reactive T^sub H^ cells are successfully generated in the presence of 1,25(OH)^sub 2^D^sub 3^, secrete proinflammatory cytokines, and do not preferentially differentiate into suppressor T cells. These cells are able to leave the lymph node, enter the peripheral circulation, and migrate to the s.c. immunization sites. However, T^sub H^ cells from 1,25(OH)^sub 2^D^sub 3^-treated mice are unable to enter the CNS parenchyma but are instead maintained in the periphery. Upon treatment cessation, mice rapidly develop experimental autoimmune encephalomyelitis, demonstrating that 1,25(OH)^sub 2^D^sub 3^ prevents the disease only temporarily likely by halting T^sub H^ cell migration into the CNS. PUBLICATION ABSTRACT
Pharmacologic targeting of T helper (T H ) cell trafficking poses an attractive opportunity for amelioration of autoimmune diseases such as multiple sclerosis (MS). MS risk is associated with vitamin ...D deficiency, and its bioactive form, 1,25-dihydroxyvitamin D3 1,25(OH) 2 D 3 , has been shown to prevent experimental autoimmune encephalomyelitis, a mouse model of MS, via an incompletely understood mechanism. Herein, we systematically examined 1,25 (OH) 2 D 3 effects on T H cells during their migration from the lymph nodes to the CNS. Our data demonstrate that myelin-reactive T H cells are successfully generated in the presence of 1,25(OH) 2 D 3 , secrete proinflammatory cytokines, and do not preferentially differentiate into suppressor T cells. These cells are able to leave the lymph node, enter the peripheral circulation, and migrate to the s. c. immunization sites. However, T H cells from 1,25(OH) 2 D 3 -treated mice are unable to enter the CNS parenchyma but are instead maintained in the periphery. Upon treatment cessation, mice rapidly develop experimental autoimmune encephalomyelitis, demonstrating that 1,25(OH) 2 D 3 prevents the disease only temporarily likely by halting T H cell migration into the CNS.
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