VXX-401, a novel anti-PCSK9 vaccine, reduces LDL-C in cynomolgus monkeys.

Atherosclerotic cardiovascular disease (ASCVD) remains the leading cause of disease burden in the world and is highly correlated with chronic elevations of LDL-C. LDL-C-lowering drugs, such as statins or monoclonal antibodies against proprotein convertase subtilisin/kexin type 9 (PCSK9), are known to reduce the risk of cardiovascular diseases; however, statins are associated with limited efficacy and poor adherence to treatment, whereas PCSK9 inhibitors are only prescribed to a "high-risk" patient population or those who have failed other therapies. Based on the proven efficacy and safety profile of existing monoclonal antibodies, we have developed a peptide-based vaccine against PCSK9, VXX-401, as an alternative option to treat hypercholesterolemia and prevent ASCVD. VXX-401 is designed to trigger a safe humoral immune response against PCSK9, resulting in the production of endogenous antibodies and a subsequent 30-40% reduction in blood LDL-C. In this article, VXX-401 demonstrates robust immunogenicity and sustained serum LDL-C-lowering effects in nonhuman primates. In addition, antibodies induced by VXX-401 bind to human PCSK9 with high affinity and block the inhibitory effect of PCSK9 on LDL-C uptake in a hepatic cell model. A repeat-dose toxicity study conducted in nonhuman primates under good laboratory practices toxicity indicated a suitable safety and tolerability profile, with injection site reactions being the main findings. As a promising safe and effective LDL-C-lowering therapy, VXX-401 may represent a broadly accessible and convenient option to treat hypercholesterolemia and prevent ASCVD.

Corneal virulence of Pseudomonas aeruginosa elastase B and alkaline protease produced by Pseudomonas putida.

PURPOSE: To measure the specific virulence contributions of two Pseudomonas aeruginosa proteases, elastase B and alkaline protease, when expressed separately by Pseudomonas putida in a rabbit model of bacterial keratitis. METHODS: P. putida KT2440 was transformed with plasmids that enabled the extracellular production of either elastase or alkaline protease. Protease expression was confirmed by zymography and immunoblotting. P. putida expressing elastase, alkaline protease, or vector alone was injected intrastromally (10(3) colony forming units [CFU]) into rabbit corneas (n=6). Infected eyes were graded by slit-lamp examination (SLE) at 20, 24, 28, and 32 hr postinfection (PI). Rabbits were sacrificed at 33 hr PI, and the log CFU (+/-SEM) per cornea was determined. RESULTS: SLE scores for eyes infected with P. putida producing elastase were significantly higher than those infected with vector alone at all time points (por=0.1), but small erosions formed in 33% of corneas. At both 24 and 28 hr PI, the SLE scores for corneas infected with P. putida producing elastase were significantly higher than those infected with P. putida producing alkaline protease (p

A humanized monoclonal antibody neutralizes yellow fever virus strain 17D-204 in vitro but does not protect a mouse model from disease.

The yellow fever virus (YFV) vaccine 17D-204 is considered safe and effective, yet rare severe adverse events (SAEs), some resulting in death, have been documented following vaccination. Individuals exhibiting post-vaccinal SAEs are ideal candidates for antiviral monoclonal antibody (MAb) therapy; the time until appearance of clinical signs post-exposure is usually short and patients are quickly hospitalized. We previously developed a murine-human chimeric monoclonal antibody (cMAb), 2C9-cIgG, reactive with both virulent YFV and 17D-204, and demonstrated its ability to prevent and treat YF disease in both AG129 mouse and hamster models of infection. To counteract possible selection of 17D-204 variants that escape neutralization by treatment with a single MAb (2C9-cIgG), we developed a second cMAb, 864-cIgG, for use in combination with 2C9-cIgG in post-vaccinal therapy. MAb 864-cIgG recognizes/neutralizes only YFV 17D-204 vaccine substrain and binds to domain III (DIII) of the viral envelope protein, which is different from the YFV type-specific binding site of 2C9-cIgG in DII. Although it neutralized 17D-204 in vitro, administration of 864-cIgG had no protective capacity in the interferon receptor-deficient AG129 mouse model of 17D-204 infection. The data presented here show that although DIII-specific 864-cIgG neutralizes virus infectivity in vitro, it does not have the ability to abrogate disease in vivo. Therefore, combination of 864-cIgG with 2C9-cIgG for treatment of YF vaccination SAEs does not appear to provide an improvement on 2C9-cIgG therapy alone.

Identification of a novel secreted protease from Pseudomonas aeruginosa that causes corneal erosions.

PURPOSE: The purpose of this study was to identify a new Pseudomonas protease and determine its possible role in keratitis. METHODS: Concentrated culture supernatants of the Pseudomonas aeruginosa strains PA103 and ATCC 19660 were analyzed by zymography. P. aeruginosa small protease (PASP) was purified from strain PA103, and modified elastase B (LasB) was purified from strain ATCC 19660. SDS-PAGE and Western blot analysis were performed on purified PASP and modified LasB. PASP was further analyzed by mass spectrometry and amino-terminal sequencing. The Pasp gene was cloned and expressed, affinity-purified in denatured form from inclusion bodies, and refolded by removal of the denaturant. Purified recombinant PASP was analyzed by zymography for protease activity. PASP and heat-inactivated PASP were injected into rabbit corneas, and the corneas were monitored for erosions caused by protease activity. RESULTS: Each strain produced a protease with a molecular mass of 80 kDa on zymograms. LasB antiserum identified the ATCC 19660 protease as modified LasB. Mass spectrometry defined the PA103 protease as having a molecular mass of 18.5 kDa. Amino-terminal sequencing and analysis of the P. aeruginosa genome sequence determined that the PA103 Pasp gene sequence was >99% identical with the PA0423 sequence of strain PAO1. Recombinant PASP was proteolytic, with a zymogram mass of 50 kDa. PASP purified from PA103 produced extensive corneal epithelial erosions, whereas heat-inactivated PASP produced no erosions. CONCLUSIONS: PASP is a protease that has not been previously identified. It causes corneal epithelial erosions, indicating its likely activity as a virulence-promoting factor in Pseudomonas keratitis.

Pathogenesis of Staphylococcus in the rabbit anterior chamber.

PURPOSE: To investigate the host defense against Staphylococcus in the rabbit anterior chamber. METHODS: The bactericidal activity of rabbit aqueous humor was investigated in vitro. Rabbit anterior chambers were injected with viable Staphylococcus aureus or Staphylococcus epidermidis (1,000 or 500,000 colony-forming units [CFU]), killed bacteria, culture supernatants of either organism, or purified S. aureus alpha-toxin. CFU as well as phospholipase (PLA(2)) and myeloperoxidase (MPO) activities of aqueous humor were determined up to 25 hours postinfection (PI). RESULTS: The number of viable S. aureus or S. epidermidis was significantly reduced when incubated with aqueous humor for 30 minutes (P

Pseudomonas aeruginosa protease IV: a corneal virulence factor of low immunogenicity.

PURPOSE: To study antibody production to Pseudomonas aeruginosa protease IV (PIV) for immunoassay development and to assess the possible role of antibody in arresting corneal damage. METHODS: Rabbits were immunized with PIV, urea-soluble recombinant PIV (rPIV), or precipitated rPIV. Antibody was analyzed by ELISA and Western blotting. Antibody-mediated inhibition of PIV activity was tested by colorimetric assay and during keratitis by slit-lamp examination of infected eyes. RESULTS: Antibody was not produced after PIV immunization but was induced by rPIV. Rabbits immunized first with soluble and then precipitated rPIV produced high titers (log(10)) to rPIV (4.28 +/- 0.09) and significantly higher titers to PIV (3.90 +/- 0.06) compared to the other immunized groups. Antibody to rPIV reacted with PIV, but neither neutralized enzyme activity in vitro nor protected infected rabbits in vivo. CONCLUSIONS: The present study demonstrates that PIV is a virulence factor which can escape a protective immune response.

Molecular analysis of Pseudomonas aeruginosa protease IV expressed in Pseudomonas putida.

PURPOSE: In this study, the protease IV gene of Pseudomonas aeruginosa was expressed in the nonocular pathogenic host, Pseudomonas putida, to elucidate the molecular properties and virulence contribution of the enzyme. Recent determination of the protease IV gene sequence suggests that the protein of 463 amino acids contains a signal sequence, a propeptide domain, and a mature protease. The only form of this protein that has been detected previously is the extracellular mature protease. METHODS: The protease IV gene was cloned and expressed in a protease IV-negative Pseudomonas species, P. putida. The cloned protease IV gene product was analyzed to identify biochemical, enzymatic, and immunologic properties and its contribution to corneal virulence. RESULTS: P. putida expressing the cloned protease IV gene had significantly greater extracellular enzyme activity than P. aeruginosa. These P. putida cell extracts produced a protein with the same molecular mass as mature protease IV and two other polypeptides representing larger precursors, all of which were recognized by protease IV-specific antibodies. P. putida producing protease IV, relative to P. putida with the vector alone, caused a threefold increase in ocular inflammation and tissue damage when intrastromally injected into rabbit corneas. CONCLUSIONS: The present study demonstrates for the first time that protease IV is synthesized as a large precursor that is processed intracellularly through an intermediate form and secreted into the extracellular milieu as a mature protease. The results also confirm a significant correlation between production of protease IV and corneal virulence.

Immunity to lysostaphin and its therapeutic value for ocular MRSA infections in the rabbit.

PURPOSE: To determine the effects of immunization against lysostaphin on the bactericidal action of lysostaphin in ocular tissue and the possible induction of allergic reactions. METHODS: Rabbits were immunized against lysostaphin by subcutaneous, intranasal, or topical routes. Anti-lysostaphin antibody titers were determined by ELISA and by neutralization of lysostaphin. Methicillin-resistant Staphylococcus aureus was intrastromally or intravitreously injected into rabbit eyes. Eyes were treated either topically with drops of lysostaphin (0.3%) or with a single intravitreous injection (0.1 mL) of lysostaphin (0.1%). At the time of death, corneas or vitreous humors were cultured to determine the number of colony forming units (CFU). RESULTS: Rabbits in keratitis experiments that were immunized subcutaneously, intranasally, or topically had serum antibody titers of 10,240, 187, and 1,867, respectively, and neutralization titers of 8 or less. In both normal and immunized rabbits with keratitis, lysostaphin significantly reduced the log CFU to less than 1 log, whereas the untreated eyes contained more than 10(6) CFU/cornea (P < or = 0.0001). Rabbits that were subcutaneously or topically immunized for endophthalmitis experiments had serum antibody titers of 1636 or 137, respectively, and neutralization titers of 2 or less. A single intravitreous injection of lysostaphin (0.1%) sterilized all eyes of immunized and nonimmune rabbits with endophthalmitis. No adverse effects were observed with the administration of lysostaphin to either normal or immunized rabbit eyes. CONCLUSIONS: Lysostaphin treatment of immunized rabbits was effective in treating S. aureus-infected eyes, despite the presence of anti-lysostaphin antibody. No adverse reactions were produced by administration of lysostaphin to immunized rabbits.

Corneal virulence of Staphylococcus aureus in an experimental model of keratitis.

The aim of this study was to determine the pathogenic role of alpha-, beta-, and gamma-toxins in a rabbit model of Staphylococcus aureus keratitis. S. aureus strains 8325-4, Newman, and their isogenic mutants were intrastromally injected into rabbit corneas. Eyes were scored for pathology by slit lamp examination (SLE), histologic examination, and bacterial colony-forming units (CFU) per cornea were determined. Rabbits were immunized against alpha-toxin and subsequently challenged with S. aureus strain 8325-4 or Newman. All strains grew equivalently to approximately 7 log CFU/cornea at 25 h postinfection. SLE scores at 15, 20, and 25 h postinfection revealed that alpha-toxin - producing strains caused greater corneal pathology than strains deficient in alpha-toxin. A beta-toxin - deficient mutant produced significantly less ocular edema than its parent or rescued strains. The gamma-toxin-deficient mutant, relative to its parent strain or genetically rescued strain, had reduced virulence. These results demonstrate that the virulence of S. aureus involves mainly alpha-toxin and to a lesser extent gamma-toxin, with beta-toxin mediating minimal corneal pathology.

Effectiveness of mupirocin and polymyxin B in experimental Staphylococcus aureus, Pseudomonas aeruginosa, and Serratia marcescens keratitis.

PURPOSE: The purpose of this study was to determine the effectiveness of mupirocin and polymyxin B, alone and in combination, in vitro and in vivo using rabbit models of, and keratitis. METHODS: Rabbit eyes were intrastromally injected with 1,000 colony-forming units (CFUs) of or or 100 CFUs of Rabbits were then treated with 2.7 mg/mL mupirocin, 10,000 U/mL polymyxin B, a mupirocin:polymyxin B combination, or 0.3% ciprofloxacin. Vehicle and untreated controls were also included. Treatment schedules depended on the strain injected. The number of CFUs was determined for all eyes after treatment. RESULTS: The mupirocin:polymyxin B combination was effective for all three genera both in vitro and in vivo. For keratitis, the mupirocin:polymyxin B combination was more effective than either drug alone and significantly reduced the log number of bacteria in the cornea by more than 3 logs compared with the vehicle or untreated controls (p

Quantitative comparison of fluoroquinolone therapies of experimental gram-negative bacterial keratitis.

PURPOSE: To determine the effectiveness of topically applied fluoroquinolones for experimental Pseudomonas or Serratia keratitis. METHODS: Bacteria were injected intrastromally (10(3) colony forming units [CFU]). From 16 to 22 hours post-infection (PI), a single topical drop of moxifloxacin (Vigamox, 0.545%), levofloxacin (Quixin, 0.5%), ofloxacin (Ocuflox, 0.3%) or ciprofloxacin (Ciloxan, 0.3%) was applied every 30 minutes. At 23 hours PI, corneas were cultured quantitatively. RESULTS: For Pseudomonas keratitis, untreated eyes contained 7 log CFU/cornea and antibiotic-treated eyes demonstrated a > or = 5-log reduction in CFU/cornea (p < or = 0.0001). Moxifloxacin, levofloxacin, or ciprofloxacin therapies were not significantly different from each other (p > or = 0.67). For Serratia keratitis, untreated eyes contained 7 logCFU/cornea whereas treated eyes had a > or = 2-log reduction (p < or = 0.0001). Moxifloxacin therapy proved most effective (p < or = 0.001). CONCLUSIONS: Overall, moxifloxacin was the most effective of the four fluoroquinolones in reducing CFU/cornea in the rabbit model of gram-negative keratitis.

Humanized monoclonal antibody 2C9-cIgG has enhanced efficacy for yellow fever prophylaxis and therapy in an immunocompetent animal model.

Yellow fever virus (YFV) causes significant human disease and mortality in tropical regions of South and Central America and Africa, despite the availability of an effective vaccine. No specific therapy for YF is available. We previously showed that the humanized monoclonal antibody (MAb) 2C9-cIgG provided prophylactic and therapeutic protection from mortality in interferon receptor-deficient strain AG129 mice challenged with YF 17D-204 vaccine. In this study we tested the prophylactic and therapeutic efficacy of this MAb against virulent YFV infection in an immunocompetent hamster model. Intraperitoneal (ip) administration of a single dose of MAb 2C9-cIgG 24h prior to YFV challenge resulted in significantly improved survival rates in animals treated with 380 or 38 µg of MAb compared to untreated animals. Treatment with the higher dose also resulted in significantly improved weight gain and reductions in serum alanine aminotransferase (ALT) and virus titers in serum and liver. Prophylactic treatment with 2C9-cIgG 24h prior to virus challenge prevented the development of a virus-neutralizing antibody (vnAb) response in hamsters. Administration of a single ip dose of 380 µg of 2C9-cIgG as late as 72 h post-YFV challenge also resulted in significant improvement in survival rates. Hamsters treated at 4-72 h post-virus challenge developed a robust vnAb response. Enhanced survival and improvement of various disease parameters in the hamster model when MAb 2C9-cIgG is administered up to 3 days after virus challenge demonstrate the clinical potential of specific antibody therapy for YF.

A small animal peripheral challenge model of yellow fever using interferon-receptor deficient mice and the 17D-204 vaccine strain.

Yellow fever virus (YFV), a member of the genus Flavivirus, is a mosquito-borne pathogen that requires wild-type (wt), virulent strains to be handled at biosafety level (BSL) 3, with HEPA-filtration of room air exhaust (BSL3+). YFV is found in tropical regions of Africa and South America and causes severe hepatic disease and death in humans. Despite the availability of effective vaccines (17D-204 or 17DD), YFV is still responsible for an estimated 200,000 cases of illness and 30,000 deaths annually. Besides vaccination, there are no other prophylactic or therapeutic strategies approved for use in human YF. Current small animal models of YF require either intra-cranial inoculation of YF vaccine to establish infection, or use of wt strains (e.g., Asibi) in order to achieve pathology. We have developed and characterized a BSL2, adult mouse peripheral challenge model for YFV infection in mice lacking receptors for interferons α, ß, and γ (strain AG129). Intraperitoneal challenge of AG129 mice with 17D-204 is a uniformly lethal in a dose-dependent manner, and 17D-204-infected AG129 mice exhibit high viral titers in both brain and liver suggesting this infection is both neurotropic and viscerotropic. Furthermore the use of a mouse model permitted the construction of a 59-biomarker multi-analyte profile (MAP) using samples of brain, liver, and serum taken at multiple time points over the course of infection. This MAP serves as a baseline for evaluating novel therapeutics and their effect on disease progression. Changes (4-fold or greater) in serum and tissue levels of pro- and anti-inflammatory mediators as well as other factors associated with tissue damage were noted in AG129 mice infected with 17D-204 as compared to mock-infected control animals.

A humanized IgG but not IgM antibody is effective in prophylaxis and therapy of yellow fever infection in an AG129/17D-204 peripheral challenge mouse model.

Yellow fever virus (YFV), a member of the genus Flavivirus, is a mosquito-borne virus found in tropical regions of Africa and South America that causes severe hepatic disease and death in humans. Despite the availability of effective vaccines, YFV is responsible for an estimated 200,000 cases and 30,000 deaths annually. There are currently no prophylactic or therapeutic strategies approved for use in human YFV infections. Furthermore, implementation of YFV 17D-204 vaccination campaigns has become problematic due to an increase in reported post-vaccinal adverse events. We have created human/murine chimeric MAbs of a YFV-reactive murine monoclonal antibody (mMAb), 2C9, that was previously shown to protect mice from lethal YFV infection and to have therapeutic activity. The new chimeric (cMAbs) were constructed by fusion of the m2C9 IgG gene variable regions with the constant regions of human IgG and IgM and expressed in Sp2 murine myelomas. The 2C9 cMAbs (2C9-cIgG and 2C9-cIgM) reacted with 17D-204 vaccine strain in an enzyme-linked immunosorbent assay and neutralized virus in vitro similarly to the parent m2C9. Both m2C9 and 2C9-cIgG when administered prophylactically 24h prior to infection protected AG129 mice from peripheral 17D-204 challenge at antibody concentrations ≥1.27 µg/mouse; however, the 2C9-cIgM did not protect even at a dose of 127 µg/mouse. The 17D-204 infection of AG129 mice is otherwise uniformly lethal. While the m2C9 was shown previously to be therapeutically effective in YFV-infected BALB/c mice at day 4 post-infection, the m2C9 and 2C9-cIgG demonstrated therapeutic activity only when administered 1 day post-infection in 17D-204-infected AG129 mice.

Development of a human-murine chimeric immunoglobulin M for use in the serological detection of human alphavirus antibodies.

Diagnosis of human alphaviral infections relies on serological techniques, such as the immunoglobulin M antibody capture-enzyme-linked immunosorbent assay (MAC-ELISA). We have humanized the alphavirus broadly cross-reactive murine monoclonal antibody 1A4B-6 to create a reagent capable of replacing human positive sera in the MAC-ELISA for diagnosis of human alphaviral infections.

Development of human-murine chimeric immunoglobulin G for use in the serological detection of human flavivirus and alphavirus antibodies.

Diagnosis of human arboviral infections relies heavily on serological techniques such as the immunoglobulin M (IgM) antibody capture enzyme-linked immunosorbent assay (MAC-ELISA) and the indirect IgG ELISA. Broad application of these assays is hindered by the lack of standardized positive human control sera that react with a wide variety of flaviviruses (e.g., dengue, West Nile, yellow fever, Japanese encephalitis, Saint Louis encephalitis, and Powassan viruses), or alphaviruses (e.g., Eastern equine encephalitis, Western equine encephalitis, Venezuelan equine encephalitis, and chikungunya viruses) that can cause human disease. We have created human-murine chimeric monoclonal antibodies (cMAbs) by combining the variable regions of flavivirus (6B6C-1) or alphavirus (1A4B-6) broadly cross-reactive murine MAbs (mMAbs) with the constant region of human IgG1. These cMAbs may be used as standardized reagents capable of replacing human infection-immune-positive control sera in indirect IgG ELISA for diagnosis of all human flaviviral or alphaviral infections. The IgG cMAbs secreted from plasmid-transformed Sp2/0-Ag14 cells had serological activity identical to that of the parent mMAbs, as measured by ELISA using multiple flaviviruses or alphaviruses.

Development of a human-murine chimeric immunoglobulin M antibody for use in the serological detection of human flavivirus antibodies.

Current diagnosis of human flaviviral infections relies heavily on serological techniques such as the immunoglobulin M (IgM) antibody capture enzyme-linked immunosorbent assay (MAC-ELISA). Broad application of this assay is hindered by a lack of standardized human positive-control sera that react with the wide variety of flaviviruses that can cause human disease, e.g., dengue virus (DENV), West Nile virus (WNV), yellow fever virus (YFV), Japanese encephalitis virus (JEV), and St. Louis encephalitis virus (SLEV). We have created a human-murine chimeric antibody combining the variable regions of the broadly flavivirus cross-reactive murine monoclonal antibody (MAb) 6B6C-1 and the constant region of human IgM to produce a standardized reagent capable of replacing human positive-control sera in a MAC-ELISA for the diagnosis of all human flaviviral infections. The human-murine chimeric IgM antibody secreted from plasmid-transformed Sp2/0-Ag14 cells had a level of serological activity identical to that of 6B6C-1 as measured by ELISA, immunoblotting, and MAC-ELISA for multiple members of the flavivirus genus, including WNV, SLEV, YFV, DENV, and JEV.

Calcium and magnesium enhance the production of Pseudomonas aeruginosa protease IV, a corneal virulence factor.

The effect of calcium and magnesium on protease IV production during the growth of Pseudomonas aeruginosa was investigated. Strain PA103 was grown to stationary phase in medium containing various concentrations of either calcium or magnesium. Culture supernatants were concentrated, standardized relative to cell density, and the pyoverdine concentrations were measured. Overall extracellular protease activity and specific protease IV (lysine endoproteinase) activity were measured with or without TLCK, a serine protease inhibitor effective against protease IV activity. Protease IV activity was also observed by casein zymography. Calcium and magnesium were quantified in the corneas and aqueous humor of rabbits that were inoculated intrastromally with strain PA103. Pyoverdine production was not significantly different in cultures grown in medium with added calcium or magnesium, but extracellular caseinase activity increased in these cultures. Susceptibility of caseinase activity to TLCK inhibition and a specific assay for protease IV indicated that protease IV activity increased in cultures grown in calcium or magnesium. Casein zymography supported the observation that protease IV activity increased in the cultures with added calcium and magnesium. Addition of calcium or magnesium to the protease IV-specific assay had no effect on the catalytic activity of pure protease IV. Infection of rabbit corneas with PA103 did not change the magnesium concentration in either corneas or aqueous humor, but significantly increased the concentration of calcium in corneas. These results indicate that calcium and magnesium enhance the production of protease IV, but not pyoverdine production. Calcium increases in the cornea following infection with P. aeruginosa could favor production of protease IV.

Pseudomonas aeruginosa protease IV degrades surfactant proteins and inhibits surfactant host defense and biophysical functions.

Pulmonary surfactant has two distinct functions within the lung: reduction of surface tension at the air-liquid interface and participation in innate host defense. Both functions are dependent on surfactant-associated proteins. Pseudomonas aeruginosa is primarily responsible for respiratory dysfunction and death in cystic fibrosis patients and is also a leading pathogen in nosocomial pneumonia. P. aeruginosa secretes a number of proteases that contribute to its virulence. We hypothesized that P. aeruginosa protease IV degrades surfactant proteins and results in a reduction in pulmonary surfactant host defense and biophysical functions. Protease IV was isolated from cultured supernatant of P. aeruginosa by gel chromatography. Incubation of cell-free bronchoalveolar lavage fluid with protease IV resulted in degradation of surfactant proteins (SP)-A, -D, and -B. SPs were degraded in a time- and dose-dependent fashion by protease IV, and degradation was inhibited by the trypsin-like serine protease inhibitor Nalpha-p-tosyl-L-lysine-chloromethyl ketone (TLCK). Degradation by protease IV inhibited SP-A- and SP-D-mediated bacterial aggregation and uptake by macrophages. Surfactant treated with protease IV was unable to reduce surface tension as effectively as untreated surfactant, and this effect was inhibited by TLCK. We speculate that protease IV may be an important contributing factor to the development and propagation of acute lung injury associated with P. aeruginosa via loss of surfactant function within the lung.

Comparison of bacterial inoculation and transcutaneous oxygen tension in the rabbit S1 perforator and latissimus dorsi musculocutaneous flaps.

Muscle and musculocutaneous flaps have been used reliably in reconstruction of soft-tissue defects for many years. Previous experimental studies have shown musculocutaneous flaps to be superior to the random pattern and fasciocutaneous flaps in the management of infected wounds. Over the past decade, perforator flaps have gained acceptance as alternative methods of reconstruction in the clinical setting that can decrease donor-site morbidity and hospital stay, and increase patient satisfaction. The authors theorized that perforator flaps may be able to handle infected wounds better than random pattern and fasciocutaneous flaps because their blood supply is essentially the same as many of their musculocutaneous counterparts. The goal of this study was to compare the S1 perforator-based skin flap and latissimus dorsi musculocutaneous flap in the dorsal flank of the rabbit with the introduction of bacteria to simulate both superficial and deep wound infection. Measurements of oxygen tension and regional perfusion index were performed on both types of flaps to ascertain their viability and capacity to heal. The authors found no statistical significance between latissimus dorsi musculocutaneous and S1 perforator flaps in the rabbit with respect to superficial and deep wound infections. The regional perfusion index was calculated for postoperative days 1, 2, and 4. No statistically significant difference between the two flaps using the regional perfusion index could be identified. Additionally, regional perfusion for both types of flaps was greater than 0.6, indicating that their capacity to heal wounds is similar.