Matrix metalloproteinases (MMPs) in fresh human prostate tumour tissue and organ-cultured prostate tissue: levels of collagenolytic and gelatinolytic MMPs are low, variable and different in fresh tissue versus organ-cultured tissue.

Prostate tissue was obtained from 22 radical prostatectomies (performed for clinical management of prostate carcinoma) immediately after surgery. A small piece of tissue was fixed immediately in formalin and used for routine histology while a second piece was frozen in OCT and used for immuno-histochemistry. Another small piece was used for isolation of epithelial and stromal cells. The remainder of the tissue was cut into 2 x 2 mm pieces and incubated in organ culture for 8 days. In organ culture, non-malignant, basal epithelial cells underwent a proliferative response. This was accompanied by de-differentiation of glandular structures and by migration of epithelial cells across the surface of the tissue. Erosion of the basement membrane could also be seen in places, but was not widespread. Invasion of epithelial cells into the adjacent stroma was not evident. Production of matrix metalloproteinases (MMPs) with gelatinolytic activity or collagenolytic activity was assessed in organ culture and compared to expression patterns in fresh tissue. MMP-1 (interstitial collagenase) and MMP-9 (92-kDa gelatinase B) were undetectable or low in fresh tissue specimens. Both enzymes were detected in organ culture and both increased over time. Even after 6 days, however, there was only a low level of gelatin-hydrolytic activity and no measurable collagen-hydrolytic activity. In past studies we used organ cultures of normal skin and malignant skin tumours (basal cell carcinomas) to help elucidate the role of collagenolytic and gelatinolytic MMPs in epithelial cell invasion (Varani et al, 2000). Compared to MMP levels observed in skin, levels of these enzymes in prostate are low. The low level of collagenolytic and gelatinolytic MMPs in fresh prostate tissue and in organ-cultured prostate tissue may help explain why there is little tissue destruction in many primary prostate tumours and why the majority of such tumours remain confined to the prostate for extended periods.

Functional expression of NAD(P)H oxidase p47 in lung microvascular endothelial cells.

Vascular endothelial cell superoxide (O(*)(2)) has an important role in intracellular signaling, in interaction with other reactive species such as nitric oxide, and in vascular dysfunction. Little is known regarding the source and function of O(*)(2) from microvascular endothelial cells from specific tissues. Mouse lung microvascular endothelial cells stimulated with phorbol ester (PMA) or NADPH generated significant O(*)(2), which was inhibited by diphenyleneiodonium (DPI) but not by allopurinol, rotenone, indomethacin, or quinacrine. Optimal O(*)(2) generation required cytosolic as well as particulate cell fractions of cells. In parallel studies, PMA induced increased expression of the p47 component of the NAD(P)H oxidase in the particulate fraction, which was inhibited by staurosporine and calphostin. These data demonstrate that NAD(P)H oxidase is an important source of O(*)(2) generation in lung microvascular endothelial cells.

Biodegradable external tracheal stents and their use in a rabbit tracheal reconstruction model.

OBJECTIVES/HYPOTHESIS: To design and develop an external biodegradable tracheal stent for use in airway reconstructive surgery. STUDY DESIGN: Experimental model. METHODS: Biodegradable external tracheal stents were fabricated from polyglycolic acid and poly(D,L-lactide-co-glycolide) (85:15). In vitro studies were performed to analyze the mechanical properties and degradative characteristics of these stents. Then the stents were tested in vivo in an anterior tracheal reconstruction model in New Zealand white rabbits. RESULTS: The average dry modulus for the external stents was 1,800 kilopascal (kPa). All of the external stents cracked by 4 weeks in buffer solution. Significant mass loss was not appreciated until after 10 weeks in solution, but by 20 weeks the stents were nearly 100% degraded. In the in vivo portion of the study, the attrition rate for the control group was 23.1% versus 50% for the external stent group. The stridor rate was approximately 38% for both groups Of the rabbits that survived the entire 3 months of the study, the stented group, when measured by a balloon catheter method, had more patent airways than the control group, with an average stenosis of 27.8% versus 47.2%, respectively (P < .05). However, more accurate postmortem cast measurements of the internal airways did not confirm this. CONCLUSIONS: The external biodegradable tracheal stent employed in this study degraded in a predictable fashion and may provide a new method to augment surgical reconstruction of the anterior tracheal wall.

Escherichia coli cells defective for the recN gene display constitutive elevation of mutagenesis at 3,N(4)-ethenocytosine via an SOS-induced mechanism.

The Escherichia coli UVM (UV Modulation of mutagenesis) response is a DNA damage-inducible mutagenic pathway detected as significantly increased mutagenesis at 3,N4-ethenocytosine (epsilon C) lesions borne on transfected single-stranded M13 vector DNA. All major classes of DNA-damaging agents can induce UVM, and the phenomenon is independent of previously characterized mutagenic responses in E. coli. To understand this phenomenon further, we set out to identify and characterize mutants in the UVM response. Screening a mutant bank of cells defective for 1-methyl-3-nitro-1-nitrosoguanidine-inducible genes revealed that defects in the recN gene cause a constitutive elevation of mutagenesis at epsilon C residues. In contrast to normal cells that show approximately 6% mutagenesis at epsilon C lesions, but approximately 60% upon UVM induction, recN-defective strains display approximately 50% mutagenesis at epsilon C lesion sites in untreated cells. However, the recN-mediated mutagenesis response was found to require the recA gene and the umuDC genes, and could be suppressed in the presence of a plasmid harbouring the SOS transcriptional repressor LexA. These results imply that recN cells are constitutively active for SOS mutagenesis functions. The observation that epsilonC mutagenesis is enhanced in recN cells confirms previous findings that mutagenesis at epsilonC can also be independently elevated by the SOS pathway.

Use of internal bioabsorbable PLGA "finger-type" stents in a rabbit tracheal reconstruction model.

OBJECTIVES: To design and develop a biodegradable tracheal stent that can be used internally to stabilize and support surgically reconstructed airways. DESIGN: In vitro mechanical and degradative properties of 80:20 poly(D,L-lactide-co-glycolide) (PLGA) "finger-like" stents were determined. The stents were then tested in vivo in rabbits that underwent anterior patch tracheoplasties with fascia lata grafts. Comparisons were made between a control group and an internal stent group for stridor development, overall group mortality, reconstructed airway lumen size, and histological findings. SUBJECTS: Twenty-five New Zealand white rabbits. RESULTS: The average dry modulus for the internal stents was 6800 kPa. All of the internal stents cracked by 4 weeks in buffer solution. Significant mass loss was not noted in vitro until after 5 weeks in buffer solution. By 14 weeks, the stents were nearly 100% degraded. The attrition rate for the control group was 23% compared with 17% for the experimental group. The stridor rate for the control group was also higher at 38% compared with 17% for the stented group. The stented rabbits had a significantly smaller average stenosis (23%) across the entire reconstruction site than the control group (34%) (P<.05). CONCLUSION: Biodegradable PLGA stents degrade in a predictable fashion and have a statistically significant effect in augmenting anterior patch tracheoplasties with fascia lata grafts in rabbits.

Endothelial cell determinants of susceptibility to neutrophil-mediated killing.

Vascular endothelial cell injury plays an important role in the pathogenesis of inflammatory-mediated tissue injury. In the current study, we assessed injury in primary cultures of endothelial cells obtained from different sites within the same species, comparing rat dermal microvascular and rat lung microvascular endothelial cells. Dermal microvascular-derived endothelial cells were more sensitive to killing by PMA (phorbol myristate acetate)-activated human neutrophils than were endothelial cells derived from lung microvasculature. Lung endothelial cells stimulated with interferon-gamma plus lipopolysaccharide (IFNgamma + LPS) generated high levels of nitric oxide (*NO), while dermal endothelial cells stimulated with IFNgamma + LPS generated significantly lower levels of *NO. Under conditions of *NO generation (IFNgamma + LPS stimulation), or in the presence of the *NO donor, S-nitroso-N-acetyl penicillamine (SNAP), endothelial cell killing by PMA-activated neutrophils was reduced. Lung endothelial cells stimulated with PMA generated less superoxide (02*-) than dermal endothelial cells. Under conditions of *NO generation (IFNgamma + LPS stimulation), or in the presence of SNAP, O2*- release from endothelial cells was reduced. Endothelial cell-derived *NO appeared to play a significant role in attenuating the neutrophil-mediated killing. The differences in the ability of endothelial cells to generate *NO and 02*- underlies, at least in part, the differences in susceptibility of these cells to injury by activated neutrophils.

Role of matrix metalloproteinases in models of macrophage-dependent acute lung injury. Evidence for alveolar macrophage as source of proteinases.

Matrix metalloproteinases (MMPs) have been implicated in the tissue injury seen in neutrophil-dependent models of acute lung injury. However, the role of MMPs in macrophage-dependent models of lung injury is unknown. To address this issue, the macrophage-dependent immunoglobulin A immune complex-induced lung injury model and the macrophage-dependent portion of the lipopolysaccharide-induced acute lung injury model in the rat were assessed for MMP involvement and for the source of these activities. In both models, injury was inhibited by the recombinant human tissue inhibitor of metalloproteinases-2. Bronchoalveolar lavage fluids (BALFs) from injured animals in both models showed increased levels of MMPs. Characterization of MMP production by isolated lung fibroblasts, endothelial cells, type II epithelial cells, and alveolar macrophages revealed that only the macrophage had the same spectrum of MMP activity as seen in the BALF. Further, isolated alveolar macrophages from injured lungs showed evidence of in vivo activation with the release of the same spectrum of MMP activities. Together these studies show that MMPs are produced during macrophage-dependent lung injury, that these MMPs play a role in the development of the lung injury, and that the alveolar macrophage is the likely source of these MMPs.

Heterogeneity of vascular endothelial cells: differences in susceptibility to neutrophil-mediated injury.

Vascular endothelial cells are key participants in the development of inflammatory-mediated tissue injury. Not all tissues have the same response to inflammation and the role of vascular endothelial cells in this response is unclear. The objective of this study was to compare cultured primary endothelial cells derived from a variety of microvascular and macrovascular sources within the same species, rat. We developed and compared primary cultures of endothelial cells from dermis, kidney and lung microvasculature, pulmonary artery and vein, renal artery and vein, and aorta. Endothelial cells were similar in their growth characteristics, ability to take up DiI-Ac-LDL, and expression of ICAM-1, regardless of their site of origin. In contrast, susceptibility to injury by PMA-activated neutrophils differed markedly among cells from different tissues as well as between microvascular and macrovascular endothelial cells from the same tissue. This study demonstrated that while endothelial cells maintain some fundamental characteristics regardless of their origin, clear site-specific differences are evident in the sensitivity of endothelial cells to injury mediated by activated neutrophils.

Escherichia coli cells expressing a mutant glyV (glycine tRNA) gene have a UVM-constitutive phenotype: implications for mechanisms underlying the mutA or mutC mutator effect.

Transfection of M13 single-stranded viral DNA bearing a 3,N4-ethenocytosine lesion into Escherichia coli cells pretreated with UV results in a significant elevation of mutagenesis at the lesion site compared to that observed in untreated cells. This response, termed UVM, for UV modulation of mutagenesis, is induced by a variety of DNA-damaging agents and is distinct from known cellular responses to DNA damage, including the SOS response. This report describes our observation, as a part of our investigation of the UVM phenomenon, that E. coli cells bearing a mutA or mutC allele display a UVM-constitutive phenotype. These mutator alleles were recently mapped (M. M. Slupska, C. Baikalov, R. Lloyd, and J. H. Miller, Proc. Natl. Acad. Sci. USA 93:4380-4385, 1996) to the glyV (mutA) and glyW (mutC) tRNA genes. Each mutant allele was shown to arise by an identical mutation in the anticodon sequence such that the mutant tRNAs could, in principle, mistranslate aspartate codons in mRNA as glycine at a low level. Because a UVM-constitutive phenotype resulting from a mutation in a tRNA gene was unexpected, we undertook a series of experiments designed to test whether the phenotype was indeed mediated by the expression of mutant glycine tRNAs. We placed either a wild-type or a mutant glyV gene under the control of a heterologous inducible promoter on a plasmid vector. E. coli cells expressing the mutant glyV gene displayed all three of the following phenotypes: (i) missense suppression of a test allele, (ii) a mutator phenotype measured by mutation to rifampin resistance, and (iii) a UVM-constitutive phenotype. These phenotypes were not associated with cells expressing the wild-type glyV gene or with cells in which the mutant allele was present but was not transcriptionally induced. These observations provide strong support for the idea that expression of mutant tRNA can confer a mutator phenotype, including the UVM-constitutive phenotype observed in mutA and mutC cells. However, our data imply that low-level mistranslation of the epsilon subunit of polymerase III probably does not account for the observed UVM-constitutive phenotype. Our results also indicate that mutA deltarecA double mutants display a normal UVM phenotype, suggesting that the mutA effect is recA dependent. The observations reported here raise a number of intriguing questions and raise the possibility that the UVM response is mediated through transient alteration of the replication environment.

Role of mismatch repair in the Escherichia coli UVM response.

Mutagenesis at 3,N4-ethenocytosine (epsilonC), a nonpairing mutagenic lesion, is significantly enhanced in Escherichia coli cells pretreated with UV, alkylating agents, or H2O2. This effect, termed UVM (for UV modulation of mutagenesis), is distinct from known DNA damage-inducible responses, such as the SOS response, the adaptive response to alkylating agents, or the oxyR-mediated response to oxidative agents. Here, we have addressed the hypothesis that UVM results from transient depletion of a mismatch repair activity that normally acts to reduce mutagenesis. To test whether the loss of mismatch repair activities results in the predicted constitutive UVM phenotype, E. coli cells defective for methyl-directed mismatch repair, for very-short-patch repair, or for the N-glycosylase activities MutY and MutM were treated with the UVM-inducing agent 1-methyl-3-nitro-1-nitrosoguanidine, with subsequent transfection of M13 viral single-stranded DNA bearing a site-specific epsilonC lesion. Survival of the M13 DNA was measured as transfection efficiency, and mutation fixation at the lesion was characterized by multiplex sequencing technology. The results showed normal UVM induction patterns in all the repair-defective strains tested. In addition, normal UVM induction was observed in cells overexpressing MutH, MutL, or MutS. All strains displayed UVM reactivation, the term used to describe the increased survival of epsilonC-containing DNA in UVM-induced cells. Taken together, these results indicate that the UVM response is independent of known mismatch repair systems in E. coli and may thus represent a previously unrecognized misrepair or misreplication pathway.

Effect of UVM induction on mutation fixation at non-pairing and mispairing DNA lesions.

Mutation fixation at an ethenocytosine (epsilon C) residue borne on transfected M13 single-stranded DNA is significantly enhanced in response to pretreatment of Escherichia coli cells with UV, alkylating agents or hydrogen peroxide, a phenomenon that we have called UVM for UV modulation of mutagenesis. The UVM response does not require the E. coli SOS or adaptive responses, and is observed in cells defective for oxyR, an oxidative DNA damage-responsive regulatory gene. UVM may represent either a novel DNA-repair phenomenon, or an unrecognized feature of DNA replication in damaged cells that affects a specific class of non-coding DNA lesions. To explore the range of DNA lesions subject to the UVM effect, we have examined mutation fixation at 3,N4-ethenocytosine and 1,N6-ethenoadenine, as well as at O6-methylguanine (O6mG). M13 viral single-stranded DNA constructs bearing a single mutagenic lesion at a specific site were transfected into cells pretreated with UV or 1-methyl-3-nitro-1-nitroso-guanidine (MNNG). Survival of transfected viral DNA was measured as transfection efficiency, and mutagenesis at the lesion site was analysed by a quantitative multiplex sequence analysis technology. The results suggest that the UVM effect modulates mutagenesis at the two etheno lesions, but does not appear to significantly affect mutagenesis at O6mG. Because the modulation of mutagenesis is observed in cells incapable of the SOS response, these data are consistent with the notion that UVM may represent a previously unrecognized DNA damage-inducible response that affects the fidelity of DNA replication at certain mutagenic lesions in Escherichia coli.

Functional recA, lexA, umuD, umuC, polA, and polB genes are not required for the Escherichia coli UVM response.

The Escherichia coli UVM response is a recently described phenomenon in which pretreatment of cells with DNA-damaging agents such as UV or alkylating agents significantly enhances mutation fixation at a model mutagenic lesion (3,N4-ethenocytosine; epsilon C) borne on a transfected M13 single-stranded DNA genome. Since UVM is observed in delta recA cells in which SOS induction should not occur, UVM may represent a novel, SOS-independent, inducible response. Here, we have addressed two specific hypothetical mechanisms for UVM: (i) UVM results from a recA-independent pathway for the induction of SOS genes thought to play a role in induced mutagenesis, and (ii) UVM results from a polymerase switch in which M13 replication in treated cells is carried out by DNA polymerase I (or DNA polymerase II) instead of DNA polymerase III. To address these hypotheses, E. coli cells with known defects in recA, lexA, umuDC, polA, or polB were treated with UV or 1-methyl-3-nitro-1-nitrosoguanidine before transfection of M13 single-stranded DNA bearing a site-specific ethenocytosine lesion. Survival of the transfected DNA was measured as transfection efficiency, and mutagenesis at the epsilon C residue was analyzed by a quantitative multiplex DNA sequencing technology. Our results show that UVM is observable in delta recA cells, in lexA3 (noninducible SOS repressor) cells, in LexA-overproducing cells, and in delta umuDC cells. Furthermore, our data show that UVM induction occurs in the absence of detectable induction of dinD, an SOS gene. These results make it unlikely that UVM results from a recA-independent alternative induction pathway for SOS gene.

Alkylating agents induce UVM, a recA-independent inducible mutagenic phenomenon in Escherichia coli.

Noninstructive DNA damage in Escherichia coli induces SOS functions hypothesized to be required for mutagenesis and translesion DNA synthesis at noncoding DNA lesions. We have recently demonstrated that in E. coli cells incapable of SOS induction, prior UV-irradiation nevertheless strongly enhances mutagenesis at a noninstructive lesion borne on M13 DNA. Here, we address the question whether this effect, named UVM for UV modulation of mutagenesis, can be induced by other DNA damaging agents. Exponentially growing delta recA cells were pretreated with alkylating agents before transfection with M13 single-stranded DNA bearing a site-specific ethenocytosine residue. Effect of cell pretreatment on survival of the transfected DNA was determined as transfection efficiency. Mutagenesis at the ethenocytosine site in pretreated or untreated cells was analyzed by multiplex DNA sequencing, a phenotype-independent technology. Our data show that 1-methyl-3-nitro-1-nitrosoguanidine, N-nitroso-N-methylurea and dimethylsulfate, but not methyl iodide, are potent inducers of UVM. Because alkylating agents induce the adaptive response to defend against DNA alkylation, we asked if the genes constituting the adaptive response are required for UVM. Our data show that MNNG induction of UVM is independent of ada, alkA and alkB genes and define UVM as an inducible mutagenic phenomenon distinct from the E. coli adaptive and SOS responses.

UVM, an ultraviolet-inducible RecA-independent mutagenic phenomenon in Escherichia coli.

Most mutagenic DNA lesions are noninstructive in the sense that template instruction is either missing or inaccessible during DNA replication, leading to replication arrest. According to the SOS hypothesis, arrested replication induces the expression of SOS factors that force replication past stalled sites at the cost of mutagenesis. We have recently shown that prior UV irradiation of delta recA cells, in which the SOS pathway does not function, enhances mutagenesis at an ethenocytosine residue borne on a circular gapped duplex DNA vector, indicating the existence of an SOS-independent inducible mutagenic phenomenon termed UVM (UV modulation of mutagenesis). In the previous experiments, mutation fixation was expected to occur during gap-filling DNA synthesis. To test whether UVM is observable during normal replication by DNA polymerase III, we have examined mutagenesis at an epsilon C residue borne on M13 single-stranded DNA. By analyzing mutation frequency and specificity using a multiplex sequence assay, we now show that UVM is observable in UV-irradiated recA+, and in delta recA cells. These data indicate that UV irradiation induces a previously unrecognized mutagenic mechanism in Escherichia coli, and that this mechanism is manifested during gap-filling DNA synthesis as well as during normal DNA replication.

Phorbol-stimulated influx of extracellular calcium in rat pulmonary artery endothelial cells.

Stimulation of rat pulmonary artery endothelial cells (RPAEC) with phorbol 12-myristate 13-acetate (PMA) resulted in an increase in intracellular calcium ([Ca2+]i). Unlike the response to bradykinin, C5a and tumor necrosis factor-alpha (TNF-alpha) previously reported (15), the PMA-induced increase in [Ca2+]i was predominantly dependent on extracellular calcium. The PMA response paralleled the BAY K 8644-induced, extracellular calcium-dependent increase in [Ca2+]i. Pretreatment of endothelial cells with the protein kinase C inhibitor staurosporine resulted in a concentration-dependent inhibition of the increase in [Ca2+]i in response to PMA. The ability of PMA analogues to induce significant increase in [Ca2+]i paralleled their ability to induce O2- generation in neutrophils. The PMA-induced influx of extracellular Ca2+ was inhibited by the L-channel selective antagonists diltiazem, nifedipine, nicardipine, and verapamil in a dose-dependent manner. Depolarizing conditions induced by high [K+]o enhanced the calcium response to PMA. The data presented are consistent with the hypothesis that PMA-induced increases in [Ca2+]i in endothelial cells are the result of Ca2+ influx through voltage-dependent L-type Ca2+ channels.

Spontaneous injury to human umbilical vein endothelial cells increases during in vitro culture and is blocked by protein kinase activation.

BACKGROUND: The in vitro culture of human umbilical vein endothelial (Huve) cells alters sensitivity to cell injury. Late passage Huve cells in basal media have an increased susceptibility to spontaneous injury and this type of cell injury is blocked by phorbol myristate acetate (PMA). EXPERIMENTAL DESIGN: Protein kinase activation and alterations in free radical generation and defense were investigated as potential mechanisms for the increased spontaneous injury of late passage Huve cells in basal media and for the PMA-mediated protective response. RESULTS: The spontaneous injury of late passage Huve cells in basal media was blocked by PMA via a protein kinase-dependent mechanism. Huve cells cultured in vitro for several passages had increased basal particulate fraction protein kinase activity and increased activity in response to PMA treatment as compared with first passage cells. The spontaneous injury was not reduced by addition of free radical scavengers or iron chelators. The increased spontaneous injury in late passage Huve cells, and the protective effects of PMA treatment were not explained by alterations in the endogenous antioxidant defense system. PMA-mediated protection against spontaneous injury was blocked by staurosporine, a protein kinase inhibitor, but not by addition of various free radical scavengers or chelators. CONCLUSIONS: These data suggest that the cellular changes mediated by protein kinase are more important than free radical metabolism or antioxidant defense as an explanation for the increased spontaneous injury of late passage Huve cells in basal media and for the ability of PMA to block this type of spontaneous cell injury.

Superoxide responses of endothelial cells to C5a and TNF-alpha: divergent signal transduction pathways.

There is increasing evidence that endothelial cells respond to a variety of mediators. In the current studies rat pulmonary artery endothelial cells (RPAEC) responded to human recombinant C5a and tumor necrosis factor-alpha (TNF-alpha) with the generation of superoxide (O2-). RPAEC responsiveness was dependent on whether cells had been obtained from confluent or subconfluent cell monolayers. RPAEC responded to C5a and TNF-alpha in a dose-dependent manner, with increases in intracellular Ca2+ (Cai2+), formation of D-myo-inositol 1,4,5-trisphosphate [Ins(1,4,5)P3], and generation of O2-. Optimal O2- responses occurred in cells that had been pretreated with the inhibitor of superoxide dismutase (SOD), diethyldithiocarbamate, and O2- responses were allopurinol insensitive. Pertussis toxin pretreatment abolished the ability of C5a to cause increases in Ins(1,4,5)P3 and Cai2+ and formation of O2- but did not inhibit the changes in Cai2+ and formation of O2- after addition of TNF-alpha. The O2- response to C5a but not to TNF-alpha was abolished by pretreatment with the inhibitor of protein kinase C, staurosporine. These data indicate that signal transduction events in response to C5a and TNF-alpha were fundamentally different.

Positive and negative elements regulate human interleukin 3 expression.

The human interleukin 3 (IL-3) promoter is comprised of several cis-acting DNA sequences that modulate T-cell expression of IL-3. These are located within 315 nucleotides upstream of the mRNA start site. Transient expression of reporter genes linked to serially deleted sequences of the IL-3 promoter has allowed mapping of two activator sequences and an interposed repressor sequence. The proximal regulatory region is specific to IL-3 and prerequisite for efficient transcription. Its effect is enhanced by a second, more distal activating sequence consisting of an AP-1 binding site. Between the two activators lies a transcriptional silencer, which is a potent repressor in the absence of the AP-1 site. DNA-nuclear protein binding experiments demonstrate specific complex formation within each of these functional regions. Thus, both positive and negative regulatory elements appear to control expression of the human IL-3 gene in activated T cells.

Ocular surface epithelium and corneal vascularization in rabbits. I. The role of wounding.

A new model for rabbit corneal vascularization, created by making a penetrating wound in corneas with epithelium of conjunctival origin, is described. Obligate resurfacing of the cornea from conjunctival epithelium usually leads to a small, but consistent peripheral superficial corneal vascularization. Subsequent penetrating wounds elicit, in 75% of cases, a marked vascular ingrowth. Normal eyes and eyes resurfaced by peripheral corneal epithelial cells do not vascularize after such wounds. The vessels are located in the anterior corneal stroma, and the regenerated epithelium has a conjunctival appearance. Although increased hydration plays a role in this vascularization, the extent of vascularization was much greater in the presence of regenerated epithelium of conjunctival origin than in the presence of regenerated epithelium of corneal origin.