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1.
Front Bioeng Biotechnol ; 10: 855777, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35795163

RESUMO

Hematopoiesis takes place in the bone marrow and is supported by a complex cellular and molecular network in the bone marrow microenvironment. Commonly used models of the human bone marrow microenvironment include murine models and two-dimensional and three-dimensional tissue cultures. While these model systems have led to critical advances in the field, they fail to recapitulate many aspects of the human bone marrow. This has limited our understanding of human bone marrow pathophysiology and has led to deficiencies in therapy for many bone marrow pathologies such as bone marrow failure syndromes and leukemias. Therefore, we have developed a modular murine bone marrow microenvironment-on-chip using a commercially available microfluidic platform. This model includes a vascular channel separated from the bone marrow channel by a semi-porous membrane and incorporates critical components of the bone marrow microenvironment, including osteoblasts, endothelial cells, mesenchymal stem cells, and hematopoietic stem and progenitor cells. This system is capable of maintaining functional hematopoietic stem cells in vitro for at least 14 days at frequencies similar to what is found in the primary bone marrow. The modular nature of this system and its accessibility will allow for acceleration of our understanding of the bone marrow.

3.
Adv Healthc Mater ; 11(7): e2101948, 2022 04.
Artigo em Inglês | MEDLINE | ID: mdl-34994104

RESUMO

Progress in the development of salivary gland regenerative strategies is limited by poor maintenance of the secretory function of salivary gland cells (SGCs) in vitro. To reduce the precipitous loss of secretory function, a modified approach to isolate intact acinar cell clusters and intercalated ducts (AIDUCs), rather than commonly used single cell suspension, is investigated. This isolation approach yields AIDUCs that maintain many of the cell-cell and cell-matrix interactions of intact glands. Encapsulation of AIDUCs in matrix metalloproteinase (MMP)-degradable PEG hydrogels promotes self-assembly into salivary gland mimetics (SGm) with acinar-like structure. Expression of Mist1, a transcription factor associated with secretory function, is detectable throughout the in vitro culture period up to 14 days. Immunohistochemistry also confirms expression of acinar cell markers (NKCC1, PIP and AQP5), duct cell markers (K7 and K5), and myoepithelial cell markers (SMA). Robust carbachol and ATP-stimulated calcium flux is observed within the SGm for up to 14 days after encapsulation, indicating that secretory function is maintained. Though some acinar-to-ductal metaplasia is observed within SGm, it is reduced compared to previous reports. In conclusion, cell-cell interactions maintained within AIDUCs together with the hydrogel microenvironment may be a promising platform for salivary gland regenerative strategies.


Assuntos
Células Acinares , Hidrogéis , Células Acinares/metabolismo , Hidrogéis/metabolismo , Metaloproteinases da Matriz/metabolismo , Glândulas Salivares/metabolismo
5.
Commun Biol ; 4(1): 361, 2021 03 19.
Artigo em Inglês | MEDLINE | ID: mdl-33742114

RESUMO

Radiation therapy for head and neck cancers causes salivary gland dysfunction leading to permanent xerostomia. Limited progress in the discovery of new therapeutic strategies is attributed to the lack of in vitro models that mimic salivary gland function and allow high-throughput drug screening. We address this limitation by combining engineered extracellular matrices with microbubble (MB) array technology to develop functional tissue mimetics for mouse and human salivary glands. We demonstrate that mouse and human salivary tissues encapsulated within matrix metalloproteinase-degradable poly(ethylene glycol) hydrogels formed in MB arrays are viable, express key salivary gland markers, and exhibit polarized localization of functional proteins. The salivary gland mimetics (SGm) respond to calcium signaling agonists and secrete salivary proteins. SGm were then used to evaluate radiosensitivity and mitigation of radiation damage using a radioprotective compound. Altogether, SGm exhibit phenotypic and functional parameters of salivary glands, and provide an enabling technology for high-content/throughput drug testing.


Assuntos
Células Acinares/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos , Ensaios de Triagem em Larga Escala , Lesões por Radiação/prevenção & controle , Glândulas Salivares/efeitos dos fármacos , Análise Serial de Tecidos , Xerostomia/prevenção & controle , Células Acinares/metabolismo , Células Acinares/efeitos da radiação , Animais , Sinalização do Cálcio/efeitos dos fármacos , Células Cultivadas , Feminino , Humanos , Hidrogéis , Masculino , Camundongos Endogâmicos C57BL , Microbolhas , Pessoa de Meia-Idade , Glândula Parótida/efeitos dos fármacos , Glândula Parótida/metabolismo , Glândula Parótida/efeitos da radiação , Fenótipo , Polietilenoglicóis/química , Lesões por Radiação/etiologia , Lesões por Radiação/metabolismo , Glândulas Salivares/metabolismo , Glândulas Salivares/efeitos da radiação , Xerostomia/etiologia , Xerostomia/metabolismo
6.
Cell Tissue Res ; 380(3): 487-497, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31900666

RESUMO

Salivary gland function is severely disrupted by radiation therapy used to treat patients diagnosed with head and neck cancer and by Sjögren's syndrome. The resulting condition, which results in xerostomia or dry mouth, is due to irreversible loss of the secretory acinar cells within the major salivary glands. There are presently no treatments for the resolution of xerostomia. Cell-based approaches could be employed to repopulate acinar cells in the salivary gland but investigations into potential therapeutic strategies are limited by the challenges of maintaining and expanding acinar cells in vitro. We investigate the encapsulation of salivary gland cell aggregates within PEG hydrogels as a means of culturing secretory acinar cells. Lineage tracing was used to monitor the fate of acinar cells isolated from murine submandibular gland (SMG). Upon initial formation in vitro, SMG aggregates comprise both acinar and duct cells, with the majority cells of acinar origin. With longer culture times, acinar cells significantly decreased the expression of specific markers and activated the expression of keratins normally found in duct cells. A similar acinar-to-duct cell transition was also observed in vivo, following duct ligation injury. These results indicate that under conditions of stress (mechanical and enzymatic isolation from glands) or injury (duct ligation), salivary gland acinar cells exhibit plasticity to adopt a duct cell phenotype.


Assuntos
Células Acinares , Plasticidade Celular , Glândula Submandibular , Células Acinares/citologia , Células Acinares/patologia , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Células Cultivadas , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Regeneração , Glândula Submandibular/citologia , Glândula Submandibular/lesões , Glândula Submandibular/patologia
7.
Arch Oral Biol ; 97: 122-130, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30384153

RESUMO

OBJECTIVE: The objective of this study was to characterize the mechanism by which salivary gland cells (SGC) aggregate in vitro. DESIGN: Timelapse microscopy was utilized to analyze the process of salivary gland aggregate formation using both primary murine and human salivary gland cells. The role of cell density, proliferation, extracellular calcium, and secretory acinar cells in aggregate formation was investigated. Finally, the ability of cells isolated from irradiated glands to form aggregates was also evaluated. RESULTS: Salivary gland cell self-organization rather than proliferation was the predominant mechanism of aggregate formation in both primary mouse and human salivary gland cultures. Aggregation was found to require extracellular calcium while acinar lineage cells account for ∼80% of the total aggregate cell population. Finally, aggregation was not impaired by irradiation. CONCLUSIONS: The data reveal that aggregation occurs as a result of heterogeneous salivary gland cell self-organization rather than from stem cell proliferation and differentiation, contradicting previous dogma. These results suggest a re-evaluation of aggregate formation as a criterion defining salivary gland stem cells.


Assuntos
Células Acinares/citologia , Glândulas Salivares/citologia , Células Acinares/efeitos da radiação , Animais , Cálcio/fisiologia , Contagem de Células , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Humanos , Camundongos , Glândulas Salivares/efeitos da radiação
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