Utility of Skin Biopsy and Culture in the Diagnosis and Classification of Chronic Ulcers: A Single-Institution, Retrospective Study.

In the United States, chronic ulcers affect 6.5 million people, with a cost of ≈$20 million annually. The most common etiology of chronic ulcers in the United States is venous stasis, followed by arterial insufficiency and neuropathic ulcers. Less common causes of chronic ulcers include infection, inflammatory etiologies such as vasculitis and pyoderma gangrenosum, and neoplastic causes. Obtaining skin biopsy and tissue culture can be helpful in diagnosing unusual causes of chronic ulcers; however, there are little data on the diagnostic utility of skin biopsy in rendering a definitive diagnosis of the etiology of chronic ulcers. A retrospective study of all skin ulcers biopsied during a 10-year period at the University of Washington was undertaken. Re-excisions and surgical wounds were excluded. A total of 270 ulcer biopsy specimens were included. In 48% of cases, no specific diagnosis could be rendered histologically. 44.8% of chronic ulcers biopsied were due to atypical causes, with neoplasms (basal cell carcinoma, squamous cell carcinoma, melanoma, and cutaneous T-cell lymphoma) being the most common. Vasculitis and pyoderma gangrenosum each represented 1.5% of rendered diagnoses. Concomitant skin culture was performed in 28.9% of cases, and special stains [acid-fast bacilli, Brown and Brenn (B&B), Grocott's methenamine silver, and periodic acid-Schiff stains] were performed in 34.0%. Although more than half (49 of 78) of tissue cultures were positive, only 6.8% (12 of 175) of special stains on tissue sections were positive. We conclude that although the etiology of many ulcers cannot be determined by routine histology alone, skin biopsy of ulcers remains a critical part of the workup given that when a specific cause can be determined, atypical etiologies, including neoplasms, represent a significant proportion of chronic ulcers. Limitations of our study include referral bias. Our results also confirm the higher diagnostic yield of conventional tissue culture compared with special tissue stain biopsies of skin ulcers.

Evaluating safety and cost-effectiveness of platelets stored in additive solution (PAS-F) as a hemolysis risk mitigation strategy.

BACKGROUND: Platelet inventory constraints can result in minor ABO incompatibility and possible hemolysis. The aims of this study were to determine the reduction of isoagglutinin in titers of platelets stored in additive solution (PAS) and compare its safety, efficiency, and cost-effectiveness with full-volume and plasma-reduced platelets. STUDY DESIGN AND METHODS: Isoagglutinin titers were performed in paired whole blood donor samples and apheresis platelets collected in PAS (PAS-PLT) aliquot samples by the tube method. RESULTS: A total of 149 pairs of donor/platelet samples were tested: 75 group O, 59 group A, and 15 group B. For group O donor samples, the median anti-A IgG and IgM were 64 and 16, respectively, and the median anti-B IgG and IgM were 64 and 16, respectively. For group O PAS-PLT samples the mean anti-A IgG and IgM, and anti-B IgG and IgM were 32 and 8, and 16 and 8, respectively. For group A donor samples, the mean anti-B IgG and IgM was 8 in both cases; and both titers decreased to 2 in PAS-PLT. For group B donor samples, mean anti-A IgG and IgM was 16 in both cases; and both titers decreased to 4 in PAS-PLT. PAS-PLT demonstrated a net reduction in cost and improved efficiency when compared to plasma reduction. The use of PAS-PLT resulted in a 40% reduction of allergic transfusion reactions. CONCLUSION: The use of PAS decreases plasma isoagglutinin titers, transfusion reactions, and is cost-effective when compared to routine plasma reduction as a strategy to mitigate hemolysis risk from minor incompatible platelet transfusion.

Highly efficient differentiation of functional hepatocytes from human induced pluripotent stem cells.

Human induced pluripotent stem cells (hiPSCs) hold great potential for use in regenerative medicine, novel drug development, and disease progression/developmental studies. Here, we report highly efficient differentiation of hiPSCs toward a relatively homogeneous population of functional hepatocytes. hiPSC-derived hepatocytes (hiHs) not only showed a high expression of hepatocyte-specific proteins and liver-specific functions, but they also developed a functional biotransformation system including phase I and II metabolizing enzymes and phase III transporters. Nuclear receptors, which are critical for regulating the expression of metabolizing enzymes, were also expressed in hiHs. hiHs also responded to different compounds/inducers of cytochrome P450 as mature hepatocytes do. To follow up on this observation, we analyzed the drug metabolizing capacity of hiHs in real time using a novel ultra performance liquid chromatography-tandem mass spectrometry. We found that, like freshly isolated primary human hepatocytes, the seven major metabolic pathways of the drug bufuralol were found in hiHs. In addition, transplanted hiHs engrafted, integrated, and proliferated in livers of an immune-deficient mouse model, and secreted human albumin, indicating that hiHs also function in vivo. In conclusion, we have generated a method for the efficient generation of hepatocytes from induced pluripotent stem cells in vitro and in vivo, and it appears that the cells function similarly to primary human hepatocytes, including developing a complete metabolic function. These results represent a significant step toward using patient/disease-specific hepatocytes for cell-based therapeutics as well as for pharmacology and toxicology studies.

Epithelial mesenchymal transition and hedgehog signaling activation are associated with chemoresistance and invasion of hepatoma subpopulations.

BACKGROUND & AIMS: Our previous studies showed that CD133, EpCAM, and aldehyde dehydrogenase (ALDH) are useful markers to identify cancer stem cells (CSCs) in hepatocellular carcinoma (HCC) tissues. The present study aims to evaluate chemosensitivity and invasion capability of HCC based on CSC marker profiles, and to explore the underlying molecular mechanisms. METHODS: Hepatoma cell lines were separated into subpopulations according to CD133, EpCAM, and ALDH expression profiles. Epithelial mesenchymal transition (EMT) and hedgehog (Hh) signaling were examined to identify their links with chemoresistance and aggressive invasion. RESULTS: Well-differentiated cell lines were positive for CD133(+)/ALDH(high) and CD133(+)/EpCAM(+) at 1.5-15% and 2.3-8.3%; whereas, poorly-differentiated cells were almost all negative for these markers. FACS-enriched CD133(+)/ALDH(high) and CD133(+)/EpCAM(+) Hep3B and Huh-7 cells formed more spheroids in vitro. CD133(-)/ALDH(low) HLE cells were more resistant to cisplatin, doxorubicin or sorafenib than their positive counterparts. CD133(-)/EpCAM(-) Huh-7 cells or CD133(-)/ALDH(-) HLE cells exhibited a higher invasion rate than their positive counterparts. HLE and HLF cells acquired EMT in double negative subpopulations. Hh activity in Huh-7 CD133(-)/EpCAM(-) cells was higher than in their positive counterparts, and the inhibition of Hh activity by cyclopamine resulted in reduced cell proliferation. CONCLUSIONS: Well-differentiated CD133(+)/ALDH(high) or CD133(+)/EpCAM(+) cells appear to be a CSC/initiating subpopulation; whereas, in poorly-differentiated hepatoma cells, EMT and enhanced hedgehog signaling activity may be responsible for their chemoresistance and invasion. These findings underscore the significance of EMT and enhanced Hh signaling in liver cancer stem or initiating cells.

New approaches in the differentiation of human embryonic stem cells and induced pluripotent stem cells toward hepatocytes.

Orthotropic liver transplantation is the only established treatment for end-stage liver diseases. Utilization of hepatocyte transplantation and bio-artificial liver devices as alternative therapeutic approaches requires an unlimited source of hepatocytes. Stem cells, especially embryonic stem cells, possessing the ability to produce functional hepatocytes for clinical applications and drug development, may provide the answer to this problem. New discoveries in the mechanisms of liver development and the emergence of induced pluripotent stem cells in 2006 have provided novel insights into hepatocyte differentiation and the use of stem cells for therapeutic applications. This review is aimed towards providing scientists and physicians with the latest advancements in this rapidly progressing field.