Tissue engineering of an auricular cartilage model utilizing cultured chondrocyte-poly(L-lactide-epsilon-caprolactone) scaffolds

To determine the potential development in vivo of tissue-engineered auricular cartilage, chondrocytes from articular cartilage of bovine forelimb joints were seeded on poly(L-lactic acid-epsilon-caprolactone) copolymer scaffolds molded into the shape of a human ear. Copolymer scaffolds alone in the same shape were studied for comparison. Chondrocyte-seeded copolymer constructs and scaffolds alone were each implanted in dorsal skin flaps of athymic mice for up to 40 weeks. Retrieved specimens were examined by histological and molecular techniques. After 10 weeks of implantation, cell-seeded constructs developed cartilage as assessed by toluidine blue and safranin-O red staining; a vascular, perichondrium-like capsule enveloped these constructs; and tissue formation resembled the auricular shape molded originally. Cartilage matrix formation increased, the capsule persisted, and initial auricular configuration was maintained through implantation for 40 weeks. The presence of cartilage production was correlated with RT-PCR analysis, which showed expression of bovine-specific type II collagen and aggrecan mRNA in cell-seeded specimens at 20 and 40 weeks. Copolymer scaffolds monitored only for 40 weeks failed to develop cartilage or a defined capsule and expressed no mRNA. Extensive vascularization led to scaffold erosion, decrease in original size, and loss of contour and shape. These results demonstrate that poly(L-lactic acid-epsilon-caprolactone) copolymer seeded with articular chondrocytes supports development and maintenance of cartilage in a human ear shape over periods to 40 weeks in this implantation model.     

A radio-frequency system for in vivo pilot experiments aimed at the studies on biological effects of electromagnetic fields

An exposure system consisting of two long transversal electromagnetic (TEM) cells, operating at a frequency of 900 MHz, is presented and discussed. The set-up allows simultaneous exposure of a significant number of animals (up to 12 mice per cell) in a blind way to a uniform plane wave at a frequency of 900 MHz, for investigating possible biological effects of exposure to electromagnetic fields produced by wireless communication systems. A heating/refrigerating system has also been designed for maintaining comfortable environmental conditions within the TEM cells during experiments. An accurate dosimetric study has been performed both numerically and by means of direct measurements on phantoms and living mice. The results have shown that good homogeneity of exposure and adequate power efficiency, in terms of whole-body specific absorption rate (SAR) per 1 W of input power, are achievable for the biological target.     

The role of N-methyl-D-aspartate (NMDA) receptors in wind-up: A mathematical model

We present a mathematical model for the phenomenon of wind-up(Mendell, 1966, Exper. Neur. 16, 316–22) which occursin many neurons. We concentrate on its occurrence in the substantiagelatinosa of the dorsal horns of the spinal cord, where itis connected with certain pathological and nonpathological painstates. The model is a development of the model by Britton &Skevington (1989, J. Theor. Biol. 137, 91–105) for MeLzack& Wall's gate control theory of pain (1965, Science, NewYork, 150, 971–9; 1982, The Challenge of Pain, Penguin:Harmondsworth), modified to take account of more recent information.Its variables are the electric potentials of various cells inthe midbrain and the spinal cord. Britton & Skevington'soriginal model simulated many of the phenomena observed in acutepain in humans, but not the wind-up mechanism. This is not surprising,since this model did not include the N-methyl-D-aspartate (NMDA)receptors that are now recognized as being crucial to the phenomenon.Here we rectify this omission, and obtain good agreement betweenthe model and experimental data on wind-up. The positive feedbackthat NMDA receptors exhibit is shown to be the essential featurein producing wind-up. As an independent test of the model wesimulate a completely different experimental set-up, and obtaingood qualitative agreement with data there. Finally, we presenta prediction of the model that has yet to be tested experimentally.     

The Involvement of Dopamine in Strengthening Cortical Signals Activating NMDA Receptors in the Striatum (a hypothetical mechanism)

A possible mechanism is proposed for the enhancement/weakening of those cortical signals in the cortex-basal ganglia-thalamus-cortex neural network which induce/do not induce opening of NMDA channels in the spiny neurons of the striatum and which can be regarded as strong/weak in terms of this measure. The mechanism is based on the modulatory influences of dopamine on changes in the efficiency of corticostriatal inputs. In the absence of dopamine, relative increases in the intensity of strong (weak) cortical signals can lead to the induction of long-term potentiation (depression) of corticostriatal synapses. In this case, because of the differently directed influences on thalamic cells of signals passing via strionigral and striopallidal cells, strong signals at the output of the thalamus are weakened, while weak signals are strengthened. Activation of dopamine D₁ (D₂) receptors on strionigral (striopallidal) neurons may facilitate increases in the extent of long-term potentiation/depression (decreases in the extent of long-term potentiation/depression or induction of long-term potentiation/depression). The consequence of this is that strong signals at the output of the thalamus can be strengthened synergistically, while weak signals cab be weakened synergistically. Background cortical signals evoking tonic release of dopamine in the striatum can decrease strengthening because of weakening of the modulatory influence of dopamine on the modification of corticostriatal synapses.     

电磁脉冲辐射对猕猴凝血机制影响研究

目的:探讨电磁脉冲对猕猴凝血机制影响。方法:经高场强电磁脉冲源以6×104V/m全身辐照5只猕猴,于照前及照后1、3、7、14、28及90d下肢前静脉采血,采用AYW-8001型血凝仪检测电磁脉冲对动物血浆中TT、PT、APTT及FIB的影响。结果:动物血浆中TT于伤后1d及14～28d延长,APTT于伤后1d延长,FIB含量于伤后14d减少,PT在照射前后无明显变化。结论:电磁脉冲照射可对实验猕猴凝血机制产生影响,可通过TT、APTT延长和FIB含量减少而使机体凝血机制发生障碍。     

Adrenalectomized mice as an experimental model for the study of the biological activity of immune complexes

Local and systemic reactions to an immune complex formed in vitro can be obtained in adrenalectomized mice. The principles for the composition of the immune complexes giving the appearance of their general (toxic) action and their ability to evoke local reactions are established. Factors inhibiting the anaphylactic reaction (injection of cortisone, antihistamines, general anesthesia) weakened the reaction to the immune complex. The local reaction to the immune complex was diminished by an artificially induced fall in the blood complement level in the animals.Presented by Academician of the Academy of Medical Sciences of the USSR V. I. Ioffe.Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 82, No. 7, pp. 847–850, July, 1976.     

A mathematical model of fluid transport in the kidney

A mathematical model of the rat kidney is developed from glomerular and tubular submodels. It is assumed that all nephrons are identical, that the hydraulic pressure in the tubules obeys Hagen-Poiseuille's law, that the rate of fluid reabsorption depends on the flow rate of tubular fluid, and that the tubules are distensible. The independent variables of the model are selected to comply with experimental measurements in the hydropenic rat. The model is used to evaluate the mechanism of glomerulotubular balance: changing the mean ultrafiltration pressure in the glomerular capillaries has a substantial influence on glomerular filtration rate (GFR). A change in the rate of fluid reabsorption in the proximal tubules has a strong influence on GFR notwithstanding that the change in GFR is smaller than that in the rate of fluid reabsorption. The calculated values for the hydraulic pressure profile in the tubular system and the interstitial pressure during ureteral obstruction are in close agreement with experimental measurements. Increasing the arterial haematocrit above normal causes a substantial decrease in GFR, whilst reducing it below normal has only a small effect on GFR.     

Protective effect of intravenous immunoglobulin (IVIG) in an experimental model of pemphigus vulgaris

Uncontrolled studies have found intravenous immunoglobulin (IVIG) to be effective in the treatment of pemphigus vulgaris (PV). The aim of this study was to evaluate the role of IVIG in preventing IgG autoantibodies binding to desmoglein-3 and blister formation using a controlled experimental design. The ability of IVIG to affect the binding of IgG affinity purified from two patients with PV to desmoglein-3 in comparison to IgG from one donor, was conducted by enzyme-linked immunosorbent assay (ELISA). The specificity was confirmed by competition assay. We assessed the effect of IVIG on the induction of experimental-PV in CD1 newborn mice by subcutaneous subjection of IgG affinity purified from two patients with PV. The treatment was conducted by subcutaneous administration of IVIG together with IgG from the pemphigus patients or appropriate control. The skin of the newborns was examined 24-48 h later for blisters, and samples of the affected areas were analysed by immunohistochemistry. IVIG as a whole molecule and its F(ab)(2) portion inhibited the binding of anti-desmoglein-3 antibody to recombinant desmoglein-3 in a dose-dependent manner. The specificity was confirmed by competition assays. In-vivo, IVIG and its F(ab)(2) portion prevented blister formation in the newborn mice. Cutaneous lesions were noted only in the groups of newborn mice who were injected with IgG fractions from the PV patients. Immunopathological evaluation revealed that IVIG prevented the formation of acanthylosis with IgG deposition in the intercellular spaces. These results point to the efficacy of IVIG in the prevention of blister formation in an experimental PV model.     

Left-right asymmetry of visual evoked potentials in brain-damaged patients: A mathematical model and experimental results

The left-right asymmetry in the potential amplitude on the scalp was studied in poststroke patients by using flash visual evoked potential (VEP) and a numerical two-dimensional model of the head. The left-right asymmetry of the VEP was measured in three patients after thrombosis, in one after hemorrhage, and in one healthy subject. The numerical model used computed tomography images to define the different comparttential distribution created by a dipole source in the occipital region was solved numerically with use of a finite volume method. Left-right asymmetry was calculated with serveral values of conductivity of the damaged region. The experimental results revealed a negative asymmetry in the three patients after thrombosis (i. e., the potential amplitude over the ischemic hemisphere was smaller than that over the intact hemisphere), whereas, in the patient after hemorrhage, a positive asymmetry was found. Nonsignificant left-right asymmetry was found in the healthy subject. The numerical model revealed that the electrical conductivity of the damaged tissue has a major effect on the left-right asymmetry. Negative asymmetry, such as that found for patients after thrombosis, was obtained when the conductivity of the damaged region was greater than that of the brain, whereas positive asymmetry (hemorrhage patient) was obtained when that conductivity was smaller than that of the brain. This finding indicates that the left-right asymmetry in the scalp VEP of patients after brain damage may be a result of changes in the conductivity of the volume conductor (the ischemic region) between the source and the electrodes.     

Effects of different natural extracts in an experimental model of benign prostatic hyperplasia (BPH)

Objective and design

To characterize the impact of inflammatory process and oxidative stress in the degree of benign prostatic hyperplasia (BPH), a common condition in which chronic inflammation plays a crucial role, we investigated the effect of different plant extract preparations in an in vivo model of BPH as new therapeutic target.

Material

BPH was made in rats with daily administration of testosterone propionate (3 mg/kg) for 14 days.

Treatment

Rats were randomized into different groups to receive oral administration of plant extract preparations: Serenoa repens with selenium (SeR 28.5 mg/kg associated with Se 0.005 mg/kg), Teoside (2 mg/kg), and Puryprost (14 mg/kg containing Teoside 50% 2 mg/kg and Epilobium 12 mg/kg).

Methods

After 14 days, rats were killed and histological changes, prostate weight and apoptotic pathways were assayed.

Results

The results obtained demonstrated that the association of treatments reduced prostate weight and hyperplasia, while treatment with Puryprost demonstrated a greater trend of protection compared to the other treatments.

Conclusion

Thus, our results indicate that plant extract could be considered as new useful therapy in the treatment of BPH with particular attention on Puryprost that represents a rational approach to reduce BPH through modulation of inflammatory process and anti-oxidant process.

     

Differentiation dependent matrix formation (fibronectin and laminin) in an experimental murine rhabdomyosarcoma model

Cellular differentiation processes along with formation of extracellular matrix proteins were investigated in methylcholanthrene-induced murine rhabdomyosarcomata. We used primary tumours, allotransplants in nude mice, and the respective tumour recurrences generated by repeated incomplete surgical tumour removal. The expression of the differentiation markers desmin and myoglobin as well as the presence of fibronectin and laminin was ascertained by immunohistochemical methods. The question arises, whether or not correlations between the grade of cellular differentiation (desmin, myoglobin) and extracellular matrix formation (fibronectin, laminin) exist in tumours with striated muscle cell differentiation. The constant relations between cellular differentiation and matrix formation in original tumours also applied to allotransplants and tumour recurrences in which partially modulations of differentiation in comparison with original tumours could be recognized.     

Morphological and functional determinants of fluoxetine (Prozac)-induced pulmonary disease in an experimental model

Fluoxetine treatment effects were determined by evaluating respiratory mechanics (elastance/resistance) and exhaled nitric oxide, as well as mononuclear and polymorphonuclear cell recruitment into the lungs, in an experimental guinea pig model. Guinea pigs were divided into four groups: Fl (fluoxetine only, n=7); Fl+Sw (fluoxetine and forced swimming, n=7); Ns+Sw (normal saline and forced swimming, n=8); and Ns (normal saline only, n=8). Treated animals received oral fluoxetine (10 mg/(kg day)) for 30 consecutive days. On day 31, all animals were anesthetized and mechanically ventilated so that respiratory system elastance and resistance, as well exhaled nitric oxide, could be determined. The lungs were then excised en bloc for histological and immunohistochemical evaluation. Forced swimming induced bronchodilation in untreated animals and bronchoconstriction in fluoxetine-treated animals. Fluoxetine treatment was also associated with mononuclear infiltration (predominantly into alveolar walls) and neutrophil recruitment. In addition, levels of exhaled nitric oxide, an inflammatory marker, were higher in fluoxetine-treated animals. Swimming-induced stress also amplified mononuclear cell recruitment to the lungs. These results show that, in this experimental model, fluoxetine treatment reproduces the pathology of chronic interstitial pneumonia in humans.     

A mathematical model of the sympathetic-heart period system in the cat

The dynamic behavior of changes in heart period in response to changes in frequency of supramaximal electrical stimulation of either pre- or postganglionic cardiac sympathetic nerves in five chloralosed cats pretreated with methyl atropine was investigated using time domain techniques. The typical response to step changes in frequency of stimulation (range 1–10 Hz) was found to be a decrease in heart period which reached a steady value after a short delay and was followed, after the stimulus was removed, by a slower return to prestimulus values. No significant difference was found between the responses to pre- and postganglionic stimulation although the strength of the stimuli required to achieve the same response was greater for postganglionic stimulation. Both the observed dynamic and steadystate responses were described to a good approximation by a first-order nonlinear system. The asymmetry in the response was accounted for by including in the model a second-order chemical reaction. The observed delay was accounted for by including a pure time delay. These results suggest that models proposed by previous investigators were unnecessarily complex.     

The starling (Sturnus vulgaris) as an experimental model for staphylococcal infection of the avian foot

Inoculation of the footpad of the starling (Sturnus vulgaris) with a broth culture of Staphylococcus aureus resulted in a swelling of the foot and histological changes similar to those seen in bumblefoot in poultry and other species. In a number of cases S: aureus could be re-isolated. It is suggested that this could prove to be a useful model in the study of avian pododermatitis.     

Osteochondral repair in the rabbit model utilizing bilayered, degradable oligo(poly(ethylene glycol) fumarate) hydrogel scaffolds

In this study, hydrogel scaffolds, based on the polymer oligo(poly(ethylene glycol) fumarate) (OPF), were implanted into osteochondral defects in the rabbit model. Scaffolds consisted of two layers-a bottom, bone forming layer and a top, cartilage forming layer. Three scaffold formulations were implanted to assess how material composition and transforming growth factor-beta1 (TGF-beta1) loading affected osteochondral repair. Critical histological evaluation and scoring of the quantity and quality of tissue in the chondral and subchondral regions of defects was performed at 4 and 14 weeks. At both time points, no evidence of prolonged inflammation was observed, and healthy tissue was seen to infiltrate the defect area. The quality of this tissue improved over time with hyaline cartilage filling the chondral region and a mixture of trabecular and compact bone filling the subchondral region at 14 weeks. A promising degree of Safranin O staining and chondrocyte organization was observed in the newly formed surface tissue, while the underlying subchondral bone was completely integrated with the surrounding bone at 14 weeks. Material composition within the bottom, bone-forming layer did not appear to affect the rate of scaffold degradation or tissue filling. However, no bone upgrowth into the chondral region was observed with any scaffold formulation. TGF-beta1 loading in the top layer of scaffolds appeared to exert some therapeutic affect on tissue quality, but further studies are necessary for scaffold optimization. Yet, the excellent tissue filling and integration resulting from osteochondral implantation of these OPF-based scaffolds demonstrates their potential in cartilage repair strategies.     

A mathematical model of microbial growth including an intermediate I. Growth in batch cultures

A mathematical model of microbial growth is presented and examined which, in contrast to the well-known MONOD model, includes transitions from one cell ?bottle-neck”? to another. This is achieved by introducing an intermediate product in the model. Three variants of the model for different regulatory functions of the intermediate are considered. The results permit to describe a set of experimentally observable microbial growth curves. According to the model, the shape of the growth curves, the kinetics of substrate consumption and changes of intermediate concentration depend on culture prehistory and the nature of the intermediate regulatory function.     

beta-catenin (CTNNB1) gene amplification: a new mechanism of protein overexpression in cancer

beta-Catenin nuclear translocation is frequently observed in different types of malignancies, including gastric cancer. In gastric cancer, however, the molecular mechanisms leading to accumulation of this protein in the nucleus remain unknown. In this setting, beta-catenin (CTNNB1) mutations have been reported, but studies of mutation frequency have yielded conflicting results. Mutations or silencing of other partners of beta-catenin (i.e., APC and AXIN) are also considered rare genetic events in gastric tumorigenesis. Gene amplification is a common mechanism of activation and/or overexpression of oncogenes in gastric and other cancers. In this study, we investigated whether gene amplification is a possible mechanism of beta-catenin activation in gastric cancer by determining its presence in 49 patients with gastric cancer and two gastric-derived cell lines (KATO III and ST2957). Using fluorescence in situ hybridization, we identified beta-catenin amplification in one of the tumor samples as well as in KATO III cells. beta-Catenin immunostaining revealed nuclear translocation of the protein in both cases. In the KATO III cells, beta-catenin overexpression was confirmed by quantitative real-time PCR and Western blot analyses and beta-catenin gene amplification by Southern blot analysis and multiplex ligation probe amplification. In the KATO III cell line, no correlation was found between beta-catenin nuclear translocation and increased expression of the WNT1 target gene CCND1 (cyclin D1). Our data suggest that gene amplification is a possible mechanism of beta-catenin overexpression in cancer.     

A tissue-engineered muscle repair construct for functional restoration of an irrecoverable muscle injury in a murine model

There are no effective clinical treatments for volumetric muscle loss (VML) resulting from traumatic injury, tumor excision, or other degenerative diseases of skeletal muscle. The goal of this study was to develop and characterize a more clinically relevant tissue-engineered muscle repair (TE-MR) construct for functional restoration of a VML injury in the mouse lattissimus dorsi (LD) muscle. To this end, TE-MR constructs developed by seeding rat myoblasts on porcine bladder acellular matrix were preconditioned in a bioreactor for 1 week and implanted in nude mice at the site of a VML injury created by excising 50% of the native LD. Two months postinjury and implantation of TE-MR, maximal tetanic force was ～72% of that observed in native LD muscle. In contrast, injured LD muscles that were not repaired, or were repaired with scaffold alone, produced only ～50% of native LD muscle force after 2 months. Histological analyses of LD tissue retrieved 2 months after implantation demonstrated remodeling of the TE-MR construct as well as the presence of desmin-positive myofibers, blood vessels, and neurovascular bundles within the TE-MR construct. Overall, these encouraging initial observations document significant functional recovery within 2 months of implantation of TE-MR constructs and provide clear proof of concept for the applicability of this technology in a murine VML injury model.