Physiologische und pathophysiologische Rolle von CD24

CD24 wird in verschiedenen Zelltypen exprimiert und ist an der Regulation von Prozessen wie Differenzierung, Proliferation und Apoptose beteiligt. Darüber hinaus wurde CD24 kürzlich in verschiedenen Geweben als Marker für adulte Stamm-bzw. Vorläuferzellen identifiziert. Obwohl CD24 auf Mausbruststammzellen exprimiert wird als auch von Bedeutung für die Progression von Brustkarzinomen ist, ist die funktionelle Rolle von CD24 in der Brust allerdings noch nicht erforscht. Aus diesem Grund war ein Ziel dieser Arbeit, die Expression und Funktion von CD24 in der Mausbrustdrüse zu charakterisieren. Mittels immunhistologischer Analysen wurde gezeigt, dass die Expression von CD24 auf luminale Brustepithelzellen beschränkt ist und abhängig vom funktionellen Differenzierungsstatus des Brustepithels einer Regulation unterliegt. So ist CD24 während der Pubertät konstant exprimiert, wird mit fortschreitender Schwangerschaft herunterreguliert und ist während der Laktation nicht mehr detektierbar. Nach Abschluss der Laktationsphase wird das ursprüngliche CD24-Expressionsniveau im Brustepithel wieder hergestellt. Zur weiteren Untersuchung der Funktion von CD24 im Mausbrustepithel wurden CD24 knock-out Mäuse herangezogen. Die morphologische Analyse zeigte dabei, dass es während der Pubertät im knock-out Brustgewebe zu einer verstärkten Verzweigung bzw. erhöhten Dichte der Brustdrüsengänge kommt, was auf eine negative regulatorische Rolle von CD24 im Verzweigungsprozess hinweist. CD24 wird von Mausbruststammzellen exprimiert, die in diesem Kontext potentielle funktionelle Rolle von CD24 wurde erstmalig in dieser Arbeit untersucht. Durch Transplantation kleiner CD24 knock-out Brustgewebestücke wurde gezeigt, dass die vollständige morphologische und funktionelle Rekonstitution des Brustgewebes erzielt werden kann. Damit ist CD24 für die Funktion von Mausbruststammzellen nicht essenziell. Eine Kompensation des Verlustes von CD24 im knock-out Brustgewebe durch verstärkte Expression des CD24-Paralogs CD52 wurde nicht beobachtet. Neben der Expression in physiologischen Kontexten spielt CD24 auch eine Rolle in verschiedenen Tumoren und ist von Bedeutung für die Progression bzw. Metastasierung. Um die funktionelle Relevanz von CD24 für die Tumorprogression in vivo zu untersuchen, wurden im Rahmen dieser Arbeit verschiedene Tumortiermodelle zunächst hinsichtlich ihrer CD24-Expression charakterisiert. Durch Verpaarung eines der Tumormodelle mit CD24 knock-out Mäusen konnte gezeigt werden, dass der Verlust der CD24-Expression eine erhöhte Überlebensrate der Tiere zur Folge hat, was auf die Beteiligung von CD24 an der Tumorprogression in diesem Modell schließen lässt

Loss of ASAP1 in mice impairs adipogenic and osteogenic differentiation of mesenchymal progenitor cells through dysregulation of FAK/Src and AKT signaling

ASAP1 is a multi-domain adaptor protein that regulates cytoskeletal dynamics, receptor recycling and intracellular vesicle trafficking. Its expression is associated with poor prognosis for a variety of cancers, and promotes cell migration, invasion and metastasis. Little is known about its physiological role. In this study, we used mice with a gene-trap inactivated ASAP1 locus to study the functional role of ASAP1 in vivo, and found defects in tissues derived from mesenchymal progenitor cells. Loss of ASAP1 led to growth retardation and delayed ossification typified by enlarged hypertrophic zones in growth plates and disorganized chondro-osseous junctions. Furthermore, loss of ASAP1 led to delayed adipocyte development and reduced fat depot formation. Consistently, deletion of ASAP1 resulted in accelerated chondrogenic differentiation of mesenchymal cells in vitro, but suppressed osteo- and adipogenic differentiation. Mechanistically, we found that FAK/Src and PI3K/AKT signaling is compromised in Asap1GT/GT MEFs, leading to impaired adipogenic differentiation. Dysregulated FAK/Src and PI3K/AKT signaling is also associated with attenuated osteogenic differentiation. Together these observations suggest that ASAP1 plays a decisive role during the differentiation of mesenchymal progenitor cells

CD24 Is Not Required for Tumor Initiation and Growth in Murine Breast and Prostate Cancer Models

CD24 is a small, heavily glycosylated, GPI-linked membrane protein, whose expression has been associated with the tumorigenesis and progression of several types of cancer. Here, we studied the expression of CD24 in tumors of MMTV-PyMT, Apc1572/T+ and TRAMP genetic mouse models that spontaneously develop mammary or prostate carcinoma, respectively. We found that CD24 is expressed during tumor development in all three models. In MMTV-PyMT and Apc1572T/+ breast tumors, CD24 was strongly but heterogeneously expressed during early tumorigenesis, but decreased in more advanced stages, and accordingly was increased in poorly differentiated lesions compared with well differentiated lesions. In prostate tumors developing in TRAMP mice, CD24 expression was strong within hyperplastic lesions in comparison with non-hyperplastic regions, and heterogeneous CD24 expression was maintained in advanced prostate carcinomas. To investigate whether CD24 plays a functional role in tumorigenesis in these models, we crossed CD24 deficient mice with MMTV-PyMT, Apc1572T/+ and TRAMP mice, and assessed the influence of CD24 deficiency on tumor onset and tumor burden. We found that mice negative or positive for CD24 did not significantly differ in terms of tumor initiation and burden in the genetic tumor models tested, with the exception of Apc1572T/+ mice, in which lack of CD24 reduced the mammary tumor burden slightly but significantly. Together, our data suggest that while CD24 is distinctively expressed during the early development of murine mammary and prostate tumors, it is not essential for the formation of tumors developing in MMTV-PyMT, Apc1572T/+ and TRAMP mice

Loss of ASAP1 in mice impairs adipogenic and osteogenic differentiation of mesenchymal progenitor cells through dysregulation of FAK/Src and AKT signaling.

ASAP1 is a multi-domain adaptor protein that regulates cytoskeletal dynamics, receptor recycling and intracellular vesicle trafficking. Its expression is associated with poor prognosis for a variety of cancers, and promotes cell migration, invasion and metastasis. Little is known about its physiological role. In this study, we used mice with a gene-trap inactivated ASAP1 locus to study the functional role of ASAP1 in vivo, and found defects in tissues derived from mesenchymal progenitor cells. Loss of ASAP1 led to growth retardation and delayed ossification typified by enlarged hypertrophic zones in growth plates and disorganized chondro-osseous junctions. Furthermore, loss of ASAP1 led to delayed adipocyte development and reduced fat depot formation. Consistently, deletion of ASAP1 resulted in accelerated chondrogenic differentiation of mesenchymal cells in vitro, but suppressed osteo- and adipogenic differentiation. Mechanistically, we found that FAK/Src and PI3K/AKT signaling is compromised in Asap1GT/GT MEFs, leading to impaired adipogenic differentiation. Dysregulated FAK/Src and PI3K/AKT signaling is also associated with attenuated osteogenic differentiation. Together these observations suggest that ASAP1 plays a decisive role during the differentiation of mesenchymal progenitor cells

CD24 is expressed during MMTV-PyMT mammary tumorigenesis.

<p>To determine the expression of CD24 during tumorigenesis, female MMTV-PyMT mice were sacrificed at a variety of ages, and their mammary glands were cut into sections and stained with antibodies specific for CD24. After counterstaining with hematoxylin, the sections of 12 animals were photographed and analysed. Representative sections are shown. Scale bars indicate 100 μm. (A) Hyperplastic preneoplastic lesions and small adenomas that are either stained strongly or negative for CD24. (B) Isotype control stained serial section corresponding to A. (C) Different degrees of CD24 staining within one and the same neoplastic lesion. (D) A histopathologic analysis was performed and the intensity of the CD24 staining was evaluated. Score:—no staining; + moderate staining; ++ strong staining. A two-sided Fisher´s exact test was performed to test the null hypothesis "staining intensity is independent of histopathologic appearance". The null hypothesis was rejected based on a calculated p-value of 0.00035 (3x3 contingency table). Scoring was categorized into CD24 negative ("-") or CD24 positive ("+" or "++"), and two-sided Fisher´s exact tests and 2x2 contingency tables were used to perform pairwise comparisons of (i) "invasive well differentiated" vs. "invasive poorly differentiated" (p = 0.21), (ii) "preinvasive" vs. "invasive well differentiated" (p = 0.45) and (iii) "preinvasive" vs. "invasive poorly differentiated" (p = 0.015). (E) More advanced tumor showing weak and diffuse staining. <b>F:</b> More advanced tumor negative for CD24.</p

CD24 deficiency does not affect MMTV-PyMT mammary tumorigenesis, but reduces tumor burden in <i>Apc</i>^{<i>1572/T+</i>} mice.

<p>To study the effect of CD24 deficiency on mammary tumor initiation and growth, MMTV-PyMT and <i>Apc</i>^{<i>1572/T+</i>} mice were crossed with <i>CD24</i>^<i>-/-</i> mice. Female MMTV-PyMT and <i>Apc</i>^{<i>1572/T+</i>} mice either <i>CD24</i>^<i>-/-</i> or <i>CD24</i>^<i>+/+</i> were regularly palpated to monitor the time point of tumor onset. Following onset, tumors were measured regularly and animals were sacrificed when the tumors reached 1 cm in diameter in one dimension. Tumors and mammary tissue were then removed in total, and their mass was determined as a measure for tumor burden. Error bars indicate SE. Significance was tested using 2-tailed un-paired t tests assuming equal variance. Scale bars indicate 100 μm. (A) Age at which tumors were first detected in MMTV-PyMT mice; <i>CD24</i>^<i>+/+</i>: n = 20; <i>CD24</i>^<i>-/-</i>: n = 21. (B) Age at which tumors were first detected in <i>Apc</i>^{<i>1572/T+</i>} mice; <i>CD24</i>^<i>+/+</i>: n = 19; <i>CD24</i>^<i>-/-</i>: n = 20. (C) Age at which MMTV-PyMT mice were sacrificed; <i>CD24</i>^<i>+/+</i>: n = 20; <i>CD24</i>^<i>-/-</i>: n = 21. (D) Age at which <i>Apc</i>^{<i>1572/T+</i>} mice were sacrificed; <i>CD24</i>^<i>+/+</i>: n = 19; <i>CD24</i>^<i>-/-</i>: n = 21. (E) MMTV-PyMT tumor burden; <i>CD24</i>^<i>+/+</i>: n = 20; <i>CD24</i>^<i>-/-</i>: n = 21. <b>F:</b> <i>Apc</i>^{<i>1572/T+</i>} tumor burden; <i>CD24</i>^<i>+/+</i>: n = 19; <i>CD24</i>^<i>-/-</i>: n = 20 * p<0.05. (G), (H) Representative hematoxylin stained sections of MMTV-PyMT <i>CD24</i>^<i>+/+</i> (left panels) and MMTV-PyMT <i>CD24</i>^<i>-/-</i> (right panels) mammary tumors. Evaluation of the sections showed poorly differentiated mammary carcinoma in situ/carcinoma (G3) with additional accompanying earlier stage lesions (hyperplasia, papilloma, adenoma) present in virtually all tumors. There was no difference between <i>CD24</i>^<i>+/+</i> and <i>CD24</i>^<i>-/-</i> MMTV-PyMT mammary tumors with the exceptions that necrotic areas were slightly less frequent in <i>CD24</i>^<i>-/-</i> tumors. (I) Representative hematoxylin stained sections of <i>Apc</i>^{<i>1572/T+</i>} <i>CD24</i>^<i>+/+</i> (left panel) and <i>Apc</i>^{<i>1572/T+</i>} <i>CD24</i>^<i>-/-</i> (right panel) mammary tumors. Histopathologic evaluation of the sections showed metaplastic carcinoma with squamous cell-like differentiation (well-differentiated; G1). There was no obvious difference between <i>CD24</i>^<i>+/+</i> and <i>CD24</i>^<i>-/-</i> <i>Apc</i>^{<i>1572/T+</i>} mammary tumors.</p

CD24 is expressed in mammary tumors during <i>Apc</i>^{<i>1572/T+</i>} tumorigenesis.

<p>To determine the expression of CD24 during tumorigenesis, female <i>Apc</i>^{<i>1572/T+</i>} mice were sacrificed at a variety of ages, and their mammary glands were cut into sections and stained with antibodies specific for CD24. After counterstaining with hematoxylin, the sections of 14 animals were photographed and analysed. Representative sections are shown. Scale bars indicate 100 μm. (A) Isotype control stained serial section corresponding to (B). (B), (C) Epithelial layers surrounding the fibrillar deposits found in squamous metaplasia staining strongly for CD24. (D) Adenocarcinoma staining weakly for CD24. (E) A histopathologic analysis was performed and the intensity of the CD24 staining was evaluated. Score:—no staining; + moderate staining; ++ strong staining. A two-sided Fisher´s exact test was performed to test the null hypothesis "staining intensity is independent of histopathologic appearance". The null hypothesis was rejected based on a calculated p-value of 0.0000002 (2x2 contingency table).</p