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PubMed1987

Human fetal tissues grafted to rodent hosts: structural and functional observations of brain, adrenal and heart tissues in oculo.

The potential for growth and development of human tissue grafts was explored by transplantation to the anterior chamber of the eye of rats and mice. Tissues were obtained from therapeutic abortions, performed in the eighth to twelfth week of gestation, using a slight modification of routine vacuum aspirations. Recipients were either adult rats immunosuppressed with cyclosporin A and protected with antibiotics, or nude immunodeficient Balb C mice. Catecholamine-rich tissues such as chromaffin cells from the adrenal medulla, sympathetic ganglia, central dopamine neuroblasts from the substantia nigra, and noradrenaline neuroblasts from the locus coeruleus all survived grafting, and in many cases formed nerve fibers that invaded the host iris. Similarly, central serotonin neurons from developing raphe nuclei grafts were able to innervate host irides. Human fetal cerebellar and cerebral cortical transplants continued their development in rat host eyes. Extracellular recordings from such cerebellar and cortical grafts revealed spontaneously active cells with immature action potential waveforms. Spinal cord grafts also survived and contained substance P-immunoreactive neurons. Dorsal root ganglia were able to form nerve fibers invading the host iris, as evidenced by neurofilament immunohistochemistry. Heart tissue survived and manifested spontaneous rhythmic contractions in oculo. Both human cortex cerebri and heart tissue grafts became innervated by sympathetic adrenergic nerve fibers from the rat host iris. Thus both graft-to-host and host-to-graft neuronal connections may be established between man and rat. Taken together, these data suggest that transplantation of human fetal nervous tissues to the anterior chamber of immunosuppressed or immunodeficient rodent hosts yields a unique model system for studies of human brain development, developmental disturbances, connectivity, and the action of drugs.

Action Potentials↗
PubMed2006

Chronic schisandrin B treatment improves mitochondrial antioxidant status and tissue heat shock protein production in various tissues of young adult and middle-aged rats.

The effects of chronic schisandrin B (Sch B) treatment (10 mg/kg/dayx15) on mitochondrial antioxidant status and sensitivity to Ca2+-induced permeability transition, as well as tissue heat shock protein (Hsp)25/70 production were examined in various tissues (brain, heart, liver, skeletal muscle) of young adult and middle-aged female rats. Age-dependent impairment in mitochondrial antioxidant status, as assessed by levels/activities of antioxidant components (reduced glutathione, alpha-tocopherol, Se-glutathione peroxidase and Mn-superoxide dismutase) and the extent of reactive oxygen species generation in vitro, was observed in brain, heart, liver and skeletal muscle tissues. While tissue Hsp25 levels remained relatively unchanged with aging, the Hsp70 level was increased in both brain and heart tissues of middle-aged rats. Chronic Sch B treatment was able to enhance mitochondrial antioxidant status and the resistance to Ca2+-induced mitochondrial permeability transition in an age-independent manner in various tissues of rats. However, Hsp25 and Hsp70 levels were only increased in young adult rats. The Sch B-induced enhancement of mitochondrial protective parameters in the heart was associated with the protection against myocardial ischemia-reperfusion injury in both young adult and middle-aged rats. The results suggest that chronic Sch B treatment may be beneficial for reversing the mitochondrial changes with aging and enhancing the heat shock response.

Aging↗
PubMed2006

Supply and use of human tissue for research purposes: survey of BATB affiliated tissue banks.

This paper describes a survey undertaken to identify the extent of supply and use of human tissue in research by BATB affiliated tissue banks. Approximately one third of tissue banks registered with the BATB are currently supplying samples that are found to be unsuitable for clinical use, for research. These banks all obtain consent for research and all supply tissue for in-house research. Some tissue is transferred to other public and commercial institutions. A harmonised network approach is proposed as the way forward to meet the increasing demand for human tissue in research.

Biomedical Research↗
PubMed2005

Tissue-specific characteristics of in vivo electric gene: transfer by tissue and intravenous injection of plasmid DNA.

PURPOSE: To evaluate the tissue-specific characteristics of electric gene transfer after tissue and intravenous injection of naked plasmid DNA (pDNA). METHODS: pDNA encoding firefly luciferase was injected directly into the liver, kidney, spleen, skin and muscle, or into the tail vein of mice, and electric pulses were then applied to one of these organs. The distribution of transgene expressing cells was evaluated using pDNA encoding beta-galactosidase. RESULTS: Tissue injection of pDNA produced a significant degree of transgene expression in any tissue with the greatest amount in the liver, followed by kidney and spleen. The expression in these organs decreased quickly with time, and muscle showed the greatest expression at 7 days. Electroporation significantly increased the expression, and the expression level was comparable among the organs. Intravenous injection of pDNA followed by electroporation resulted in a significant expression in the liver, spleen, and kidney but not in the skin or muscle. CONCLUSIONS: Electric gene transfer to the liver, kidney, and spleen can be an effective approach to obtain significant amounts of transgene expression by either tissue or intravenous injection of pDNA, whereas it is only effective after tissue injection as far as skin- or muscle-targeted gene transfer is concerned.

Animals↗
PubMed1987

Immunological evidence for the presence of hormone-sensitive lipase in rat tissues other than adipose tissue.

A polyclonal rabbit antibody was used to detect hormone-sensitive lipase in rat organs other than white adipose tissue. Inhibition of tissue diacylglycerol lipase activity by the anti-hormone-sensitive lipase, and by NaF, Hg2+ and diisopropyl fluorophosphate, known inhibitors of the hormone-sensitive lipase, demonstrated its presence in the adrenals, ovaries, testes, heart and skeletal muscle, but not in the liver and kidneys. After enrichment by immunoprecipitation an immunoreactive protein, corresponding to the adipose tissue hormone-sensitive lipase 84 kDa subunit, and some additional, higher Mrapp proteins, were detected by Western blotting in the same tissues. The adipose tissue contained greater than 80% of the total hormone-sensitive lipase, with 5-10- and 50-100-fold lower specific activity in the steroid-producing and the muscle tissues, respectively.

Adipose Tissue↗
PubMed2006

Stereological methods to assess tissue response for tissue-engineered scaffolds.

The stereological approach can provide an objective unbiased assessment of structural change in biological systems. In this review, we elucidate the basic principles of stereology and their implementation in the analysis of tissue response to tissue-engineering scaffolds. A brief outline of tissue response parameters that can be estimated using stereological approach is included. The focus is on frequently quantified parameters in tissue response, such as host tissue infiltration, inflammatory cell numbers, angiogenesis, fibrous tissue thickness, areas of calcification, and/or necrosis, among others. Special consideration is given to sampling techniques and how these techniques can influence the reliability of the obtained results as well as minimizing potential sources of bias. These basic principles are illustrated with practical examples, where measurements are performed and estimations calculated using conventional stereological techniques. As the next generation of biomaterials continue to be developed, it is essential that researchers develop a rigorous and unbiased method of performance quantification.

Animals↗
PubMed2006

Extraction of RNA from archival tissues and measurement of thrombospondin-1 mRNA in normal, dysplastic, and malignant oral tissues.

Thrombospondin-1 (TSP-1) is an extracellular matrix glycoprotein implicated in the regulation of angiogenesis and tumour development. Our objectives were to ascertain the quantity and quality of RNA extracted from archival, formalin-fixed, paraffin embedded, oral tissues and their application in measuring the concentrations of TSP-1 mRNA in these tissues. We compared three techniques of isolation of RNA as well as related experimental variables. TSP-1 mRNA was measured in specimens of normal, dysplastic, and malignant oral tissues by real-time reverse transcriptase polymerase chain reaction (RT-PCR). RNA suitable for analysis by real-time RT-PCR was obtained by the three techniques tested, although the yield varied depending on the protocol used (range 0.2-3.6 microg/mm(3)). The mean (S.D.) concentrations of TSP-1 mRNA relative to 18S were 21.1 (7.2) in normal oral tissues (n=9), 11.0 (8.2) in dysplastic tissue (n=8) and 7.3 (5.3) in carcinomatous tissue (n=17). The difference between normal and carcinomatous specimens was significant (p=0.01). This reduction in expression of TSP-1 mRNA from normal to dysplasia to carcinoma may favour the angiogenic drive that accompanies the development of oral tumours.

Carcinoma, Squamous Cell↗
PubMed2006

Tissue kallikrein and kininogen levels in fetoplacental tissues from normotensive pregnant women and women with pregnancy-induced hypertension.

OBJECTIVES: An imbalance of vasoconstrictor and vasodilator substances in the placenta has been postulated in the pathogenesis of pregnancy-induced hypertension (PIH). There is however little information available on the kallikrein-kinin system (KKS) in women with PIH. The aim of this study therefore was to determine tissue kallikrein and kininogen levels and their distribution patterns in fetoplacental tissues from both normotensive pregnant (NTP) women and women with PIH. STUDY DESIGN: The study group consisted of 24 women, 12 of whom had normal pregnancies, while 12 had PIH. Portions of amnion, chorion laeve, placental plate chorion, fetal placenta and maternal placenta were dissected from each freshly delivered placenta. Tissue kallikrein (total, active and inactive) and kininogen levels were estimated using a synthetic chromogenic substrate, S-2266 and an enzyme immunoassay, respectively. The data were analysed using Mann-Whitney U and Kruskal-Wallis tests. RESULTS: No significant differences were found for total, active and inactive tissue kallikrein levels in all fetoplacental tissues between both groups. However, kininogen levels were found to be significantly lower in chorion laeve, placental plate chorion, fetal placenta and maternal placenta from women with PIH when compared to those in similar tissues from NTP women. CONCLUSION: These findings suggest the presence of an abnormality in the kallikrein-kinin system in the placentas of women with PIH, which requires further study.

Case-Control Studies↗
PubMed2006

Prevention of device-related tissue damage during percutaneous deployment of tissue-engineered heart valves.

BACKGROUND: Endovascular application of pulmonary heart valves has been recently introduced clinically. A tissue-engineering approach was pursued to overcome the current limitations of bovine jugular vein valves (degeneration and limited longevity). However, deployment of the delicate tissue-engineered valves resulted in severe tissue damage. Therefore the objective of this study was to prevent tissue damage during the folding and deployment maneuver. MATERIAL AND METHODS: Porcine pulmonary heart valves, small intestinal submucosa, and ovine carotid arteries were obtained from a slaughterhouse. After dissection and antimicrobial incubation, the valves were trimmed (removal of sinus and most of the muscular ring) to fit into the deployment catheter. The inside (in-stent group, n = 6) or outside (out-stent group, n = 6) of a nitinol stent was covered by an acellular small intestinal submucosa, and the valves were sutured into the stent. The valves were folded, tested for placement in the deployment catheter, and decellularized enzymatically. Myofibroblasts were obtained from carotid artery segments and seeded onto the scaffolds. The seeded constructs were placed in a dynamic bioreactor system and cultured for 16 consecutive days. After endothelial cell seeding, the constructs were folded, deployed, and processed for histology and surface electron microscopy. RESULTS: The valves opened and closed competently throughout the entire dynamic culture. Surface electron microscopy revealed an almost completely preserved tissue in the in-stent group. Stents covered with small intestinal submucosa on the outside, however, showed severe damage. CONCLUSION: This study demonstrates that small intestinal submucosa covering of the inside of a pulmonary valved stent can prevent stent strut-related tissue damage.

Animals↗
PubMed1997

Free tissue transfer to extend the limits of limb salvage for lower extremity tissue loss.

BACKGROUND: The extent of tissue loss amenable to primary healing after revascularization is unknown. Salvage of limbs with large soft-tissue defects with exposed tendon, joint, or bone lies beyond the limits of conventional techniques. We report our results using free tissue transfer as an adjunct to lower extremity vascular reconstruction in patients with complex ischemic or infected wounds. METHODS: Retrospective chart review of patient and wound characteristics. RESULTS: From January 1992 to June 1996, 585 procedures were performed in 544 patients, including 27 free flaps in 26 patients: 17 free flaps combined with distal bypass (7 staged, 10 simultaneous) and 10 isolated free flaps. Flap donor sites included radial forearm (8), latissimus dorsi (7), rectus abdominus (9), and scapula (3). Surgical indications included extensive ischemic/neurotrophic ulcers, and nonhealing vein graft harvest incision or transmetatarsal amputation site. Mean area of tissue loss was 70 cm2, mean ulcer duration was 5 months, and 92% of patients had exposed tendon, joint, or bone. During a mean follow-up of 14 months, 2 patients died of cardiopulmonary disease and 3 flaps failed, resulting in below-knee amputation. Six flaps were revised for graft stenosis (1), venous thrombosis (1), or flap edge necrosis (4). Limb salvage rate was 70% at 24 months by life-table analysis. Functional ambulation was achieved in 21 of 24 (88%) patients, including 7 of 8 with diabetes, end-stage renal disease, and heel ulcers. CONCLUSION: In select ambulatory patients with large soft-tissue defects and exposed deep structures, functional limb salvage is obtainable in more than 80% of patients. For lesions not amenable to vascular reconstruction with conventional methods of wound coverage, free tissue transfer extends the limits of limb salvage and is a viable alternative to amputation.

Adult↗
PubMed2002

Bladder augmentation in dogs using the tissue capsule formed around a perivesical tissue expander.

PURPOSE: Enterocystoplasty provides needed improvement in bladder storage parameters in many patients but it also generates significant morbidity. We evaluated the unusual potential alternative of using the capsule that forms around a standard silicone tissue expander placed perivesically to augment the bladder in dogs. MATERIALS AND METHODS: Six mongrel dogs underwent baseline videourodynamics and assessment of serum electrolytes, followed by placement of a 250 to 500 cc perivesical silicone tissue expander. Four months after implantation the tissue expander was removed and the fibrous capsule around the expander was biopsied. The capsule was opened and anastomosed to the bladder to augment storage. Serum electrolytes were determined 2 and 4 weeks after augmentation. Videourodynamics were repeated after 3 to 5 months, that is at sacrifice. The harvested bladders underwent histological evaluation. RESULTS: Five dogs underwent augmentation as described, while in an additional dog that underwent intraperitoneal placement of the tissue expander a fibrous capsule failed to form. Of the 5 augmented dogs 4 underwent repeat urodynamic and electrolyte evaluation with harvesting of the lower urinary tract, while 1 died of undetermined causes 3 weeks after augmentation. A distinct capsule formed in all dogs and augmentation was technically achievable. Anastomosis calcification in 3 dogs limited filling the augmenting capsule for cystography. Bladder capacity and compliance improved in all animals but it varied in degree. Histological examination of the capsule biopsies showed collagen rich connective tissue without epithelium or smooth muscle. After augmentation the capsular segment revealed urothelium in all cases with squamous metaplasia in 1. The subepithelial region had dense fibrosis and a thin band of osseous metaplasia occupied the lamina propria in all cases. Disorganized smooth muscle bundles were noted in all augmented bladders within the collagen of the capsule wall. CONCLUSIONS: Bladder augmentation with the collagenous capsule formed over a perivesical tissue expander is technically feasible. There was evidence of epithelial and smooth muscle ingrowth from the native bladder with improved bladder capacity and compliance in all dogs. Osseous metaplasia of the luminal surface of the collagen based capsule that developed in all animals may have been responsible for anastomotic narrowing and limited filling on cystography.

Animals↗
PubMed1996

Influence of tissue preparation on the high-frequency acoustic properties of normal kidney tissue.

The influence of various tissue preparations on the acoustic properties of normal kidney tissue at high frequencies was investigated. Eight surgically excised normal kidney tissue specimens were classified into three groups: (i) fresh, frozen section, (ii) formalin-fixed, frozen section and (iii) formalin-fixed, paraffin section. Scanning acoustic microscopy operating in the frequency range of 100-200 MHz was used to display the two-dimensional distribution of attenuation constant and sound speed. Our results indicate that (i) there is no significant variation in both acoustic parameters between the three tissue groups, (ii) fixation by 10% formalin produces no significant change in the acoustic parameters, (iii) in fat-free tissue regions, the acoustic parameters are independent of preparation method and (iv) frozen sections must be used to assess the acoustic parameters in fat-rich tissues.

Histocytological Preparation Techniques↗
PubMed2003

Prediction of in vivo tissue distribution from in vitro data. 3. Correlation between in vitro and in vivo tissue distribution of a homologous series of nine 5-n-alkyl-5-ethyl barbituric acids.

PURPOSE: To evaluate the ability to determine accurate in vivo tissue-to-unbound plasma distribution coefficients (Kpue) from in vitro data. METHODS: Fresh pieces of fifteen rat tissues/organs were incubated at 37 degrees C with a homologous series of nine barbiturates covering a wide range of lipophilicity (Log P 0.02 to 4.13). Steady-state in vivo Kpue values were estimated from the tissue and plasma concentrations following simultaneous dosing by constant rate i.v. infusion of all nine barbiturates. Drug concentrations in the tissues and media were determined by HPLC with UV or mass spectrometric detection. RESULTS: The pharmacokinetics of the barbiturate series following constant rate i.v. infusion indicated a range of clearance (0.49 to 30 ml x min(-1) x kg(-1)) and volume of distribution at steady state (0.51 to 1.9 l x kg(-1)) values. Good agreement was observed between the in vitro and in vivo Kpu values, although for the most lipophilic barbiturates the in vitro data underpredicted the in vivo tissue distribution for all tissues. CONCLUSION: The in vitro system for predicting the extent of in vivo tissue distribution works well for compounds of widely differing lipophilicity, although for the most lipophilic drugs it may result in an underprediction of in vivo values.

Animals↗
PubMed1999

[Direct in-vitro measurement of ultrasound velocity in carcinomas, mastopathic tissue, fatty tissue and fibroadenomas of the female breast].

PURPOSE: The study aimed to investigate ultrasound velocity (SV) in carcinomas, fibrocystic changes, fibroadenomas and fatty tissue of the female breast by means of direct in-vitro measurements. We intended to test whether or not differences in SV exist between the various types of tissue and whether the SV is a useful criterion to differentiate the different tissues. METHOD: SV was measured by comparing transmission time of the ultrasound beam through the specimen and through water. Altogether 40 specimens (12 cancer, 14 fibrocystic changes = FCD, 10 fatty tissues, 3 fibroadenomas, and 1 mixed tissue) were analysed. RESULTS: Velocity differed significantly between fat (1478.5 +/- 6.5 m/s) and tumor (1523.1 +/- 5.9 m/s) (p approximately 10(-11)) and between fat and FCD (1526.0 +/- 9.0 m/s) (p approximately 10(-12)). No significant differences and much overlap were seen between the ultrasound velocities of tumors and FCD. Ultrasound velocity in fibroadenomas (1533.2 +/- 3.8 m/s) was comparable with that in carcinomas and FCD. CONCLUSIONS: We conclude that ultrasound velocity may add complementary information to echogenicity (B-scan). Thus, a locally exact correlation of echogenicity and sound velocity might allow for an improved tissue characterization.

Adipose Tissue↗
PubMed1996

Vitamin K status in human tissues: tissue-specific accumulation of phylloquinone and menaquinone-4.

We measured the vitamin K status in postmortem human tissues (brain, heart, kidney, liver, lung, pancreas) to see if there is a tissue-specific distribution pattern. Phylloquinone (K1) was recovered in all tissues with relatively high levels in liver, heart and pancreas (medians, 10.6 (4.8), 9.3 (4.2), 28.4 (12.8) pmol(ng)/g wet weight tissue); low levels (< 2 pmol/g) were found in brain, kidney and lung. Menaquinone-4 (MK-4) was recovered from most of the tissues; its levels exceeded the K1 levels in brain and kidney (median, 2.8 ng/g) and equalled K1 in pancreas. Liver, heart and lung were low in MK-4. The higher menaquinones, MK-6-11, were recovered in the liver samples (n 6), traces of MK-6-9 were found in some of the heart and pancreas samples. The results show that in man there are tissue-specific, vitamin-K distribution patterns comparable to those in the rat. Furthermore, the accumulation of vitamin K in heart, brain and pancreas suggests a hitherto unrecognized physiological function of this vitamin.

Adult↗
PubMed2002

Tissue-engineering bioreactors: a new combined cell-seeding and perfusion system for vascular tissue engineering.

One approach to the tissue engineering of vascular structures is to develop in vitro conditions in order ultimately to fabricate functional vascular tissues before final implantation. In our experiment, we aimed to develop a new combined cell seeding and perfusion system that provides sterile conditions during cell seeding and biomechanical stimuli in order to fabricate autologous human vascular tissue in vitro. The cell seeding and perfusion system is made of Plexiglas and is completely transparent (Berlin Heart, Berlin, Germany; University Hospital Benjamin Franklin, Berlin, Germany). The whole system consists of a cell seeding chamber that can be incorporated into the perfusion system and an air-driven respirator pump connected to the bioreactor. The cell culture medium continuously circulates through a closed-loop system. We thus developed a cell seeding device for static and dynamic seeding of vascular cells onto a polymeric vascular scaffold and a closed-loop perfused bioreactor for long-term vascular conditioning. The cell seeding chamber can be easily connected to the bioreactor, which combines continuous, pulsatile perfusion and mechanical stimulation to the tissue-engineered conduit. Adjusting the stroke volume, the stroke rate, and the inspiration/expiration time of the ventilator allows various pulsatile flows and different levels of pressure. The whole system is a highly isolated cell culture setting, which provides a high level of sterility and a gas supply and fits into a standard humidified incubator. The device can be sterilized by ethylene oxide and assembled with a standard screwdriver. Our newly developed combination of a cell seeding and conditioning device provides sterile conditions and biodynamic stimuli for controlled tissue development and in vitro conditioning of an autologous tissue-engineered vessel.

Bioreactors↗
PubMed2006

Tissue-engineered lung: an in vivo and in vitro comparison of polyglycolic acid and pluronic F-127 hydrogel/somatic lung progenitor cell constructs to support tissue growth.

In this study, we describe the isolation and characterization of a population of adult-derived or somatic lung progenitor cells (SLPC) from adult mammalian lung tissue and the promotion of alveolar tissue growth by these cells (both in vitro and in vivo) after seeding onto synthetic polymer scaffolds. After extended in vitro culture, differentiating cells expressed Clara cell 10kDa protein, surfactant protein-C, and cytokeratin but did not form organized structures. When cells were combined with synthetic scaffolds, polyglycolic acid (PGA) or Pluronic F-127 (PF-127), and maintained in vitro or implanted in vivo, they expressed lung-specific markers for Clara cells, pneumocytes, and respiratory epithelium and organized into identifiable pulmonary structures (including those similar to alveoli and terminal bronchi), with evidence of smooth muscle development. Although PGA has been shown to be an excellent polymer for culture of specific cell types in vitro, in vivo culture in an immunocompetent host induced a foreign body response that altered the integrity of the developing lung tissue. Use of PF-127/cell constructs resulted in the development of tissue with less inflammatory reaction. These data suggest that the therapeutic use of engineered tissues requires both the use of specific cell phenotypes, as well as the careful selection of synthetic polymers, to facilitate the assembly of functional tissue.

Animals↗
PubMed1982

Evaluation of a tissue transport medium for immunological characterization of benign and malignant lymphoid tissues.

A tissue transport medium was evaluated for its usefulness in immunologic characterization of lymphocytes in tissue sections. A variety of lymphoid tissues were studied, including normal tissue, hyperplastic tissue and malignant lymphoma. Approximately equal parts of biopsy samples were taken and one part was immediately snap-frozen while the other was held in transport medium for up to one week before processing for frozen section. Sections were stained with fluorescein-conjugated anti-immunoglobulin antisera and with a variety of murine monoclonal antibodies to T-cell antigens or Ia antigen followed by staining with fluorescein-conjugated goat-antimouse gamma globulin. In all cases studied, there was complete agreement in the staining patterns of tissues which were either immediately snap-frozen or held in transport medium.

Antibodies, Monoclonal↗