Charles L. Dupin

Aging alters tissue resident mesenchymal stem cell properties

Tissue resident mesenchymal stem cells (MSCs) are known to participate in tissue regeneration that follows cell turnover, apoptosis, or necrosis. It has been long known that aging impedes an organisms repair/regeneration capabilities. In order to study the age associated changes, the molecular characteristics of adipose tissue derived MSCs (ASCs) from three age groups of healthy volunteers, i.e., young, middle aged, and aged were investigated. The number and multilineage differentiation potential of ASCs declined with age. Aging reduces the proliferative capacity along with increases in cellular senescence. A significant increase in quiescence of G2 and S phase was observed in ASCs from aged donors. The expression of genes related to senescence such as CHEK1 and cyclin-dependent kinase inhibitor p16(ink4a) was increased with age, however genes of apoptosis were downregulated. Further, an age-dependent abnormality in the expression of DNA break repair genes was observed. Global microRNA analysis revealed an abnormal expression of mir-27b, mir-106a, mir-199a, and let-7. In ubiquitously distributed adipose tissue (and ASCs), aging brings about important alterations, which might be critical for tissue regeneration and homeostasis. Our findings therefore provide a better understanding of the mechanism(s) involved in stem cell aging and regenerative potential, and this in turn may affect tissue repair that declines with aging.

Persistent High Glucose Concentrations Alter the Regenerative Potential of Mesenchymal Stem Cells

Type 2 diabetes is associated with numerous long-term complications. This study aims to investigate whether impaired function of tissue-resident multipotent cells play role in pathogenesis of allied complications. Adipose-tissue-derived mesenchymal stem cells (ASCs) derived from nondiabetic (nASCs) and diabetic (dASCs) donors were compared with regard to glucose metabolism, cell replication, apoptosis, and differentiation potential. The data evidenced that elevation of glucose reduces proliferative capacity of both dASCs and nASCs, but impacts dASCs more significantly. Incorporation of insulin enhanced cell replication especially in nASCs. dASCs show higher levels of cellular senescence and apoptosis than nASCs. Unlike nASCs, apoptosis is induced via intrinsic pathway in dASCs. Data also evidenced that high glucose concentrations cause prominent disparities in nASCs and dASCs in expression of genes involved in insulin resistance such as adiponectin and resistin. Some changes in gene expression were irreversible in dASCs when treated with insulin. Additionally, high glucose concentrations reduce osteogenic and chondrogenic potential of ASCs, but enhance adipogenic potential. These results indicate that in addition to involvement in insulin resistance, impaired function of mesenchymal stem cells that reside in adipose tissue as one of the major sources of adult stem cells might be responsible for complications related to diabetes type 2.

Prospective dual role of mesenchymal stem cells in breast tumor microenvironment.

Breast cancer tissue is a heterogeneous cellular milieu comprising cancer and host cells. The interaction between breast malignant and non-malignant cells takes place in breast tumor microenvironment (TM), and has a crucial role in breast cancer progression. In addition to cellular component of TM, it mainly consists of cytokines released by tumor cells. The tumor-tropic capacity of mesenchymal stem cells (MSCs) and their interaction with breast TM is an active area of investigation. In the present communication, the interplay between the breast resident adipose tissue-derived MSCs (B-ASCs) and breast TM was studied. It was found that a distinct subset of B-ASCs display a strong affinity for conditioned media (CM) from two breast cancer cell lines, MDA-MB 231 (MDA-CM) and MCF-7 (MCF-CM). The expressions of several cytokines including angiogenin, GM-CSF, IL-6, GRO-α and IL-8 in MDA-CM and MCF-CM have been identified. Upon functional analysis a crucial role for GRO-α and IL-8 in B-ASCs migration was detected. The B-ASC migration was found to be via negative regulation of RECK and enhanced expression of MMPs. Furthermore, transcriptome analysis showed that migratory subpopulation express both pro- and anti-tumorigenic genes and microRNAs (miRNA). Importantly, we observed that the migratory cells exhibit similar gene and miRNA attributes as those seen in B-ASCs of breast cancer patients. These findings are novel and suggest that in breast cancer, B-ASCs migrate to the proximity of tumor foci. Characterization of the molecular mechanisms involved in the interplay between B-ASCs and breast TM will help in understanding the probable role of B-ASCs in breast cancer development, and could pave way for anticancer therapies.

Mesenchymal stem cell derived hematopoietic cells are permissive to HIV-1 infection.

BackgroundTissue resident mesenchymal stem cells (MSCs) are multipotent, self-renewing cells known for their differentiation potential into cells of mesenchymal lineage. The ability of single cell clones isolated from adipose tissue resident MSCs (ASCs) to differentiate into cells of hematopoietic lineage has been previously demonstrated. In the present study, we investigated if the hematopoietic differentiated (HD) cells derived from ASCs could productively be infected with HIV-1.ResultsHD cells were generated by differentiating clonally expanded cultures of adherent subsets of ASCs (CD90+, CD105+, CD45-, and CD34-). Transcriptome analysis revealed that HD cells acquire a number of elements that increase their susceptibility for HIV-1 infection, including HIV-1 receptor/co-receptor and other key cellular cofactors. HIV-1 infected HD cells (HD-HIV) showed elevated p24 protein and gag and tat gene expression, implying a high and productive infection. HD-HIV cells showed decreased CD4, but significant increase in the expression of CCR5, CXCR4, Nef-associated factor HCK, and Vpu-associated factor BTRC. HIV-1 restricting factors like APOBEC3F and TRIM5 also showed up regulation. HIV-1 infection increased apoptosis and cell cycle regulatory genes in HD cells. Although undifferentiated ASCs failed to show productive infection, HIV-1 exposure increased the expression of several hematopoietic lineage associated genes such as c-Kit, MMD2, and IL-10.ConclusionsConsidering the presence of profuse amounts of ASCs in different tissues, these findings suggest the possible role that could be played by HD cells derived from ASCs in HIV-1 infection. The undifferentiated ASCs were non-permissive to HIV-1 infection; however, HIV-1 exposure increased the expression of some hematopoietic lineage related genes. The findings relate the importance of ASCs in HIV-1 research and facilitate the understanding of the disease process and management strategies.

Cheek and inferior eyelid reconstruction after skin cancer ablation

Most patients with actinic lesions and skin cancer are skin type I or II, older than 50 years of age, and have a history of extensive sunlight exposure. These patients have been treated in our units according to universal principles. A multidisciplinary team approach can produce encouraging long-term results. The size and depth of the lesion are assessed in planning the ablation. The residual defect after the tumor resection is anticipated in the preoperative plan. Adequate resection is mandatory, even if the reconstruction must be delayed to ensure clear margins. Attention to unit and subunit anatomy facilitates adequate reconstruction with acceptable deformity. Placing scars in borders or along the lines of minimal skin tension reduces deformity. Planning the flap so that the donor site is in tissue areas with maximum laxity guards against donor site deformity. Flaps must be planned to avoid excess tension on the lower lid and central face. Attempts should be made to reduce scarring in the central face as seen in the frontal view. Respecting these principles will allow for reconstruction of the largest facial unit in a manner acceptable to the patient.

Localization of the dominant deep inferior epigastric artery perforator by computed tomography angiogram: does the standard deep inferior epigastric artery perforator flap design include the dominant perforator?

BackgroundThe deep inferior epigastric artery perforator (DIEP) free flap is the optimal autogenous reconstructive technique in many patients undergoing postmastectomy. Our aim was to evaluate the standard DIEP free flap design in relation to the dominant perforating vessels using computed tomography angiography (CTA). MethodsWe retrospectively reviewed CTAs from 75 patients who had undergone perforator flap reconstruction within the past year. Locations of the largest perforator with a minimum diameter of 2.0 mm piercing the anterior rectus fascia were recorded. ResultsOf 150 hemiabdomens reviewed, 146 (97.3%) had a dominant perforator. The median location for the dominant perforator was 3.3 cm lateral and 0.9 cm below the umbilicus. One hundred twenty-one (83%) of the dominant perforators arose within 3 cm of the umbilicus. One hundred one (69%) arose at or below the level of the umbilicus. Forty-five (31%) arose above the level of the umbilicus. Thirteen (9%) arose more than 2 cm above the umbilicus. ConclusionsThe standard DIEP flap design incorporates most of the dominant perforating vessels. However, a significant number of perforators arise at or above the umbilicus, which would be near the edge or out of the standard design of the DIEP. Our findings support the use of preoperative CTA in the evaluation of patients undergoing DIEP free flap reconstruction. Modification of flap design to include the dominant perforating vessels should be considered when the dominant vessel is outside the standard design of the DIEP.

Composite fronto-orbital reconstruction using a subscapular chimeric flap.

Composite defects of the fronto-orbital region represent a reconstructive challenge. Total autogenous reconstruction requires a thin pliable skin flap along with a bony component that can be osteotomized predictably. The thoracodorsal artery perforator/scapular bone chimeric flap provides a reliable single-stage method to cover difficult composite forehead defects where local options are unsuitable or have been exhausted. We present a patient with a traumatic fronto-orbital defect that has been successfully reconstructed using this flap. The report of this case illustrates well the versatility of the subscapular system as a donor site for difficult composite posttraumatic defects where tissue components need to be inset in a complex three-dimensional fashion.

Revision Breast Reconstruction and General Complications

Virtually all procedures for breast reconstruction require some form of revision for optimal outcome. Revision surgery is normally done as an outpatient. Implant and expander reconstructions require removal of the temporary implant and placement of a permanent prosthetic. Autologous procedures normally have a second procedure for reconstructing the nipple-areolar complex and other related matters. The general complications of reconstruction are discussed as well as advice about how to avoid and treat complications.

Autologous Breast Reconstruction

The progress in the last 25 years, utilizing the patient’s own tissue for reconstruction, has been amazing. Starting with the landmark procedure of the transverse rectus abdominis flap by Carl Hartrampf, autologous reconstruction has become much more reliable. The aesthetic outcomes have improved as well. This chapter discusses the various types of flaps which are currently employed and their advantages and disadvantages.

A History of Breast Reconstruction Following Mastectomy

Breast reconstruction has had a period of rapid development during the past 25 years. In the late 1970s, the reliability and quality of breast reduction following mastectomy were very suboptimal. During the quarter of a century, reconstruction has become very reliable and has a very high acceptance rate among patients.