High-resolution optical coherence tomography as a non-destructive monitoring tool for the engineering of skin equivalents

BACKGROUND: Three dimensional skin equivalents are widely used in dermatopharmacological and toxicological studies and as autologous transplants in wound healing. In pharmacology, there is tremendous need for monitoring the response of engineered skin equivalents to external treatment. Transplantation of skin equivalents for wound healing requires careful verification of their quality prior to transplantation. Optical coherence tomography (OCT) is a non-contact, non-destructive imaging technique for living tissues offering the potential to fulfill these needs. This work presents an analysis of OCT for high-resolution monitoring of skin equivalents at different stages during the culture process. METHODS: We developed a high-resolution OCT imaging setup based on a commercially available OCT system. A broadband femtosecond laser light source replaces the original superluminescence diode. Tomograms of living skin equivalents were recorded with an axial resolution of 3 mum and correlated with histology and immunofluorescence images. Comparison with standard low-resolution OCT is presented to emphasize the advantages of high-resolution OCT for this application. RESULTS: OCT is particularly able to distinguish between different layers of skin equivalents including stratum corneum, epidermal and dermal layer as well as the basement membrane zone. The high-resolution OCT scans correlate closely with two key benchmarks, histology and immunofluorescence imaging. CONCLUSIONS: This study clearly demonstrates the benefits of high-resolution OCT for identifying living tissue structure and morphology. Compared with the current gold standard histology, OCT offers non-destructive tissue imaging, enabling high-resolution evaluation of living tissue morphology and structure as it evolves.     

Neuronal regeneration after application of radiofrequency energy to collagenous tissue is affected by limb immobilization: an in vivo animal study.

Despite widespread use of radiofrequency (RF)-shrinkage, there have been no studies on the influence of RF-energy on neural elements of collagenous tissue. The purpose of this study was to examine the effect of RF-shrinkage on neural structures of capsuloligamentous tissue and the recovery of neural elements under different postoperative treatment protocols. One patellar tendon of 46 New-Zealand-White rabbits was shrunk. Six rabbits were sacrificed immediately postoperative. Twenty rabbits were not immobilized, 10 were immobilized for 3 and 10 were immobilized for 6 weeks. A monoclonal antibody, specific against a neurofilament protein, was used to detect nerves and neural structures. Staining pattern of nerve fibres was significantly altered immediately postoperative. After 3 weeks the number of nerve fibres and bundles decreased significantly in immobilized and non-immobilized limbs. The loss of nerve fibres was significantly less in immobilized limbs. At 6 weeks the number of neural elements in immobilized limbs increased to the level of untreated control tissue. In non-immobilized limbs we found no recovery of neural elements 9 weeks postoperatively. At this time the number of nerve fibres and bundles was still significantly less compared to the untreated control limbs. RF-shrinkage causes significant alteration of neural elements. Under immobilization nerve fibres and bundles reach the level of normal untreated tissue. Careful rehabilitation is important after RF-shrinkage. Not only for biomechanical reasons, but also to allow the neural elements to recover, thermally modified tissue should be protected from normal physiologic loads.     

Ocular hemodynamics following suction cup oculopression

In the study reported here the authors investigated the influence on ocular perfusion pressures of suction-cup oculopression, a procedure which permits oculopression without orbital compression. Patients over 50 years of age were examined before cataract extraction. Ulrich's method of ocular oscillodynamography was used to determine ocular perfusion pressures. A Mikuni and Yoneyama suction-cup dynamometer was applied for suction-cup oculopression. Ocular hypotonia following suction-cup oculopression was adequate and comparable to that achieved by other oculopression methods. After oculopression a significant increase in ocular perfusion pressures (retinal, ciliary, and diastolic) was found. However, ocular blood pressure remains constant after oculopression. Moreover, the increase in ocular perfusion pressure could not be explained by oculocardiac reflexes, because there was no change in systemic arterial blood pressure. It must be assumed, therefore, that the increase in ocular perfusion pressures is caused by a lowering of resistance, i.e., a drop in intraocular pressure.     

Antagonism of airway reactivity induced by ovalbumin antigen in guinea pigs by 5-amino-4-imidazolecarboxamide riboside

The effect of 5-amino-4-imidazolecarboxamide riboside (AICA riboside), a modulator of purine metabolism, was studied on antigen-induced bronchospasm in ovalbumin (OA)-sensitized guinea pigs. In separate experiments, sodium cromoglycate (SCG) and terbutaline were used to compare their effectiveness with that of AICA riboside (wt/vol). AICA riboside and SCG were administered as an aerosol daily for a minimum of 2 weeks before OA aerosol challenge. Terbutaline, as an aerosol, was administered once 5 minutes before OA challenge. Airway reactivity was determined through the use of a whole-body plethysmography by monitoring specific airway resistance (SRaw). OA aerosol challenge of 0.05%, 0.1%, and 0.25% (wt/vol), administered for a period of 1 minute, increased SRaw. Each of the three agents attenuated the effect of OA on SRaw, although terbutaline demonstrated more consistency and potency as compared to either AICA riboside or SCG. However, at moderate degrees of OA challenge, AICA riboside appeared to be as effective as either agent. Although the mechanism of action of AICA riboside remains uncertain, it may have therapeutic benefit in the treatment of asthma or allergic diseases.     

Detection and localization of single-base sequence differences in foot-and-mouth disease virus genomes by the RNase mismatch cleavage method.

The RNase mismatch cleavage method was examined for its efficiency of indicating single-base sequence differences in the capsid protein-coding regions of different foot-and-mouth disease virus subtype O1 strains. The method was found suitable for indicating such differences. RNase A as well as RNase T1 contributed to substrate conversion. Examples for the cleavage of eleven different single-base mismatches in RNA double-strands are now known. All virus genomes found to differ from each other exhibited three or more non-neighboured single-base sequence differences. Other genomes found to be indistinguishable by this method were those of a recent field isolate adapted to cell culture, and those of a vaccine production strain; its progeny was transmitted to pig and cow and then analyzed. The results suggest that host change does not necessarily select for antigenic variant virus, and that virus submitted to some kind of selection pressure is changed at more than one position.     

Pulmonale Thrombendarterektomie: Die Therapie der chronisch thromboembolischen pulmonalen Hypertension— Initiale chirurgische Erfahrung

Zusammenfassung Grundlagen Die pulmonale Thrombendarterektomie stellt eine effektive Therapie zur Behandlung von Patienten mit chronisch thromboembolischer pulmonalar Hypertonie dar. Wir berichten über unsere Erfahrungen mit dieser Operationstechnik bei den ersten 9 Patienten. Methodik Zwischen 1992 und Oktober 1994 wurde bei 9 Patienten eine pulmonale Thrombendarterektomie durchgeführt. über eine mediane Sternotomie wurden die Pulmonalarterien beider Seiten bis in ihre subsegmentalen Aufzweigungen im Sinne einer echten Endarterektomie vom organisierten thromboembolischen Material befreit. Zumeist waren Perioden des totalen Kreislaufstillstandes in tiefer Hypothermie notwendig, wobei diese zunehmend verkürzt werden konnten und 3 Patienten vollst?ndig ohne Kreislaufstopp operiert wurden. Ergebnisse Die perioperative Mortalit?t betrug 11%, wobei der 1. Patient am 14. postoperativen Tag an den Folgen eines Reperfusions?dems verstarb. Alle anderen Patienten sind 4 bis 22 Monate (im Mittel 13 Monate) nach dem Eingriff am Leben und zeigen eine Hochsignifikante Verbesserung ihrer h?modynamischen Situation (mean PAP pr?operativ: 62 mm Hg, postoperativ: 30 mm Hg p<0,001; Cardiac Index pr?operativ: 2,1 1/min/m², postoperativ: 3,6 l/min/m² p=0,001). W?hrend pr?operativ alle Patienten in NYHA-Klasse III oder IV waren, sind die 8 überlebenden jetzt alle in Klasse I oder II. Schlu?folgerungen Die pulmonale Thrombendarterektomie bietet eine erfolgreiche Therapiem?glichkeit für Patienten mit chronisch thromboembolischer pulmonaler Hypertonie mit einer in Anbetracht fehlender konservativer Altermativen akzeptablen Mortalit?t und ausgezeichneten funktionellen Langzeitergebnissen.      

Physiological incompatibilities of porcine hepatocytes for clinical liver support.

In fulminant hepatic failure, the use of bioartificial liver support (BAL) with porcine hepatocytes is the subject of a current and controversial debate.1 Specifically, the issue of cross-species physiological incompatibilities has not been addressed so far. We therefore investigated the effects of species-specific cytokines in single and cocultures on hepatocyte function. Hepatocyte cultures were isolated from human resection specimens and from Landrace pigs. Single and cocultures were exposed to porcine and human interleukin (IL)-6 or tumor necrosis factor (TNF)-alpha. Changes in expression of C-reactive protein (CRP), albumin, CCAAT enhancer binding protein (C/EBP)-alpha and C/EBP-beta and metabolic competence of cultured cells was studied by measuring testosterone metabolite production. After human or porcine IL-6 dosing, CRP was induced up to 100-fold in human hepatocyte cultures, while porcine hepatocytes responded marginally (2- to 5-fold). Treatment with human or porcine IL-6 or TNF-alpha resulted in reduced albumin production, albeit at different levels when human and porcine hepatocytes were compared (P = 0.001). Unlike human, porcine hepatocytes produced less of 6alpha-hydroxytestosterone (6alpha-HT) (P < 0.001) and 7alpha-HT (P < 0.001) after human or porcine IL-6 dosing and treatment with species-specific TNF-alpha induced (human hepatocytes) or decreased (porcine hepatocytes) 6beta-HT production (P = 0.021). In coculture with free exchange of metabolites, porcine hepatocytes produced less 6alpha-HT (P = 0.048) and 16alpha-HT (P = 0.033), whereas after treatment with human IL-6 reduced CRP gene and protein expression was observed with human hepatocytes (P = 0.013). In conclusion, species-specific responses of hepatocytes to cytokines and interactions with xenobiotic metabolites may limit the clinical effectiveness of porcine hepatocytes in BAL.