The combined effect of oxidative stress and TRPV1 on temperature-induced asthma: Evidence in a mouse model.

Temperature is one of the possible activators for asthma. As global warming continues, the health hazard of high temperatures is increasing. It is unclear, nevertheless, how high temperatures affect asthma. The research aims to examine how asthma is affected by high temperatures and underlying molecular mechanisms. The BALB/c mice were adopted in a model of asthma. The mice were exposed at 24 °C, 38 °C and 40 °C for 4h on weekdays from day 1 to day 30. After the experiment, the lung function was measured in vivo, and then serum protein, pulmonary inflammation and immunohistochemistry assay was assessed in vitro. As the temperature increased from 24 °C to 40 °C, there was a significant increase in serum protein, while there is no discernible difference in serum protein of OVA-sIgE and OVA-sIgG between the OVA (38 °C) group and OVA (24 °C) group. The immunohistochemistry assay showed a change in the pro-inflammatory cytokines. The histopathological analysis exhibited the change of airway structure after high-temperature exposure, especially for exposure at 40 °C. The results of signals protein showed a remarkable rise of TRPV1 for OVA+40 °C. Our results revealed that high temperatures may make asthmatic airway dysfunction severe, and the higher the temperature, the more serious asthma. The oxidative stress and TRPV1 receptor can be a potential drug target for asthma. It will provide a new tool for precision medicine in asthma.

Chemical "Butterfly Effect" Explaining the Coordination Chemistry and Antimicrobial Properties of Clavanin Complexes.

Can a minor difference in the nonmetal binding sequence of antimicrobial clavanins explain the drastic change in the coordination environment and antimicrobial efficiency? This study answers the question with a definite "yes", showing the details of the bioinorganic chemistry of Zn(II) and Cu(II) complexes with clavanins, histidine-rich, antimicrobial peptides from hemocytes of the tunicate Styela clava.

Occurrence of Spontaneous Cortical Spreading Depression Is Increased by Blood Constituents and Impairs Neurological Recovery after Subdural Hematoma in Rats.

Acute subdural hemorrhage (ASDH) is common and associated with severe morbidity and mortality. To date, the role of spontaneous cortical spreading depression (sCSD) in exaggerating secondary injury after ASDH, is poorly understood. The present study contains two experimental groups: First, we investigated and characterized the occurrence of sCSD after subdural blood infusion (300 µL) via tissue impedance (IMP) measurement in a rat model. Second, we compared the occurrence and influence of sCSD on lesion growth and neurological deficit in the presence and absence of whole blood constituents. In the first experimental group, three IMP traits could be distinguished after ASDH: no sCSD, recurrent sCSD, and constant elevated IMP (anoxic depolarization [AD]). In the second experimental group, sCSD occurred more often after autologous blood, compared with paraffin oil infusion. Lesion volume 7 days post-ASDH was 27.3 ± 6.8 mm3 after blood and 3.4 ± 2.1 mm3 after paraffin oil infusion. Subgroup analysis showed larger lesion size in animals with sCSD, than in those without. Further, occurrence of sCSD led to worse neurological outcomes in both groups. sCSD occurs early after ASDH and does not depend on the presence of whole blood constituents. However, numbers and degree of sCSD are more frequent and severe after autologous blood infusion, compared with an inert volume substance. The occurrence of sCSD leads to lesion growth and worse neurological outcome. Thus, our data advocate close monitoring and targeted treatment of sCSD after ASDH evacuation.

Characterization of Two Novel Lipopolysaccharide Phosphoethanolamine Transferases in Pasteurella multocida and Their Role in Resistance to Cathelicidin-2.

The lipopolysaccharide (LPS) produced by the Gram-negative bacterial pathogen Pasteurella multocida has phosphoethanolamine (PEtn) residues attached to lipid A, 3-deoxy-d-manno-octulosonic acid (Kdo), heptose, and galactose. In this report, we show that PEtn is transferred to lipid A by the P. multocida EptA homologue, PetL, and is transferred to galactose by a novel PEtn transferase that is unique to P. multocida called PetG. Transcriptomic analyses indicated that petL expression was positively regulated by the global regulator Fis and negatively regulated by an Hfq-dependent small RNA. Importantly, we have identified a novel PEtn transferase called PetK that is responsible for PEtn addition to the single Kdo molecule (Kdo1), directly linked to lipid A in the P. multocida glycoform A LPS. In vitro assays showed that the presence of a functional petL and petK, and therefore the presence of PEtn on lipid A and Kdo1, was essential for resistance to the cationic, antimicrobial peptide cathelicidin-2. The importance of PEtn on Kdo1 and the identification of the transferase responsible for this addition have not previously been shown. Phylogenetic analysis revealed that PetK is the first representative of a new family of predicted PEtn transferases. The PetK family consists of uncharacterized proteins from a range of Gram-negative bacteria that produce LPS glycoforms with only one Kdo molecule, including pathogenic species within the genera Vibrio, Bordetella, and Haemophilus We predict that many of these bacteria will require the addition of PEtn to Kdo for maximum protection against host antimicrobial peptides.

Cytotoxicity associated with prolonged room temperature storage of serum and proposed methods for reduction of cytotoxicity.

Canine serum preserved at room temperature (25°C) for longer than 24h is known to exhibit significant cytotoxicity. This phenomenon is one of the major reasons for the failure of virus neutralization tests. In this study, a method for reducing this cytotoxicity was investigated by applying several treatments to dog, cat and human serum prior to room temperature storage. Additionally, the identity of the cytotoxic factor generated during room temperature storage was investigated. Heat-inactivation at 56°C or 65°C and the addition of protease inhibitor prior to storage were found to be effective for reducing cytotoxicity in the serum. Furthermore, heat-inactivation at 65°C reduced the cytotoxicity that was induced under room temperature storage. Several protein factors in serum were suspected to play a role in the observed cytotoxicity. According to this study, the membrane-attack-complex in serum was not involved in the cytotoxicity. This study provides useful information for development and improvement of cell culture and virus neutralization tests.

Regulatory T cell-derived IL-10 ameliorates crescentic GN.

Regulatory T cells (Tregs) exert their immunosuppressive activity through several immunoregulatory mechanisms, including the production of anti-inflammatory cytokines such as IL-10. Although several studies suggest a role for Tregs in modulating crescentic GN, the underlying mechanisms are not well understood. Here, using IL-10 reporter mice, we detected IL-10-producing Foxp3(+) T cells in the kidney, blood, and secondary lymphoid tissue in a mouse model of crescentic GN. Specific inactivation of Il10 in Foxp3(+) Tregs eliminated the ability of these cells to suppress renal and systemic production of IFNγ and IL-17; these IL-10-deficient Tregs lost their capacity to attenuate renal tissue injury. These data highlight the suppressive functions of Tregs in crescentic GN and suggest the importance of Treg-derived IL-10 in ameliorating disease severity and in modulating both the Th1 and most notably Th17 immune response.

In vitro toxicity of serum protein-adsorbed citrate-reduced gold nanoparticles in human lung adenocarcinoma cells.

We examined the cytotoxicity effect of the serum protein coated gold nanoparticles (AuNPs) in the A549 cells. Negatively charged AuNPs were prepared by chemical reduction using citrate. The dimension and surface charge of AuNPs were characterized using transmission electron microscopy (TEM), dynamic light scattering (DLS), and zeta potential measurements. The AuNPs modified by the citrate anion were presumed to adsorb the serum proteins as indicated from the visible absorption spectroscopy, DLS, and quartz crystal microbalance (QCM) data. The QCM results indicated that among the constituents, fetal bovine serum (FBS) should be the major adsorbate species on the AuNPs incubated in the RPMI medium. The internalization of AuNPs into the A549 cells was also monitored using TEM and dark-field microscopy (DFM). Both methylthiazol tetrazolium (MTT) and lactate dehydrogenase (LDH) assays revealed that AuNPs were toxic as determined by their half-maximal inhibitory concentration. A flow cytometric and real-time PCR analysis of apoptotic genes along with the ATP depletion measurements suggested that AuNPs induce cell damages through extrinsic and intrinsic apoptotic pathways.

Sera from amyotrophic lateral sclerosis patients induce the non-canonical activation of NMDA receptors "in vitro".

Amyotrophic lateral sclerosis (ALS) is a neuromuscular disease characterized by the selective loss of both upper and lower motoneurons (MNs). The familial form of the illness is associated with mutations in the gene encoding Cu/Zn superoxide dismutase 1 (SOD-1) enzyme, but it accounts for fewer than 10% of cases; the rest, more than 90%, correspond to the sporadic form of ALS. Although many proposals have been suggested over the years, the mechanisms underlying the characteristic selective killing of MN in ALS remain unknown. In this study we tested the effect of sera from sporadic ALS patients on NMDA receptors (NMDAR). We hypothesize that an endogenous seric factor is implicated in neuronal death in ALS, mediated by the modulation of NMDAR. Sera from ALS patients and from healthy subjects were pretreated to inactivate complement pathways and dialyzed to remove glutamate and glycine. IgGs from ALS patients and healthy subjects were obtained by affinity chromatography and dialyzed against phosphate-buffered saline. Human NMDAR were expressed in Xenopus laevis oocytes, and ionic currents were recorded using the two-electrode voltage clamp technique. Sera from sporadic ALS patients induced transient oscillatory currents in oocytes expressing NMDAR with a significantly higher total electrical charge than that induced by sera from healthy subjects. Sera from patients with other neuromuscular diseases did not exert this effect. The currents were inhibited by MK-801, a noncompetitive blocker of NMDAR. The PLC inhibitor, U-73122, and the IP(3) receptor antagonist, 2-APB, also inhibited the sera-induced currents. The oscillatory signal recorded was due to internal calcium mobilization. Isolated IgGs from ALS patients significantly affected the activity of oocytes injected with NMDAR, causing a 2-fold increase over the response recorded for IgGs from healthy subjects. Our data support the notion that ALS sera contain soluble factors that mobilize intracellular calcium, not opening directly the ionic conductance, but through the non-canonical activation of NMDAR.

Analogues of peptide SMAP-29 with comparable antimicrobial potency and reduced cytotoxicity.

SMAP-29 (sheep myeloid antimicrobial peptide-29) is a peptide with potent antibacterial properties. However, it is also highly cytotoxic both to human red blood cells (hRBCs) and human embryonic kidney (HEK) cells. In this study, some of the amino acids of SMAP-29 were changed in an attempt to reduce haemolytic activity whilst maintaining high antibacterial efficacy. These analogues, plus other analogues described in the literature with potent antimicrobial activity against Gram-positive bacteria coupled with no or low haemolytic activity, were evaluated for their cytotoxicity (hRBCs and HEK cells) as well as antimicrobial efficacy against two Gram-positive (Bacillus anthracis and Bacillus globigii) and two Gram-negative bacteria (Escherichia coli and Burkholderia thailandensis). The analogues previously described in the literature were found to have low antibacterial and haemolytic activity. Two of the designed analogues had comparable antibacterial efficacy with SMAP-29 against B. anthracis but reduced haemolytic activity and therefore had a therapeutic index that was enhanced 2.3-2.6-fold over that of SMAP-29.

Effects of plasma from patients with acute on chronic liver failure on function of cytochrome P450 in immortalized human hepatocytes.

BACKGROUND: The bioartificial liver is anticipated to be a promising alternative choice for patients with liver failure. Toxic substances which accumulate in the patients' plasma exert deleterious effects on hepatocytes in the bioreactor, and potentially reduce the efficacy of bioartificial liver devices. This study was designed to investigate the effects of plasma from patients with acute on chronic liver failure (AoCLF) on immortalized human hepatocytes in terms of cytochrome P450 gene expression, drug metabolism activity and detoxification capability. METHODS: Immortalized human hepatocytes (HepLi-2 cells) were cultured in medium containing fetal calf serum or human plasma from three patients with AoCLF. The cytochrome P450 (CYP3A5, CYP2E1, CYP3A4) expression, drug metabolism activity and detoxification capability of HepLi-2 cells were assessed by RT-PCR, lidocaine clearance and ammonia elimination assay. RESULTS: After incubation in medium containing AoCLF plasma for 24 hours, the cytochrome P450 mRNA expression of HepLi-2 cells was not significantly decreased compared with control culture. Ammonia elimination and lidocaine clearance assay showed that the ability of ammonia removal and drug metabolism remained stable. CONCLUSIONS: Immortalized human hepatocytes can be exposed to AoCLF plasma for at least 24 hours with no significant reduction in the function of cytochrome P450. HepLi-2 cells appear to be effective in metabolism and detoxification and can be potentially used in the development of bioartificial liver.

Characterisation and evaluation of synthetic antimicrobial peptides against Bacillus globigii, Bacillus anthracis and Burkholderia thailandensis.

Antimicrobial peptides (AMPs) are produced by all forms of living organisms and represent a novel class of antibiotics to treat infectious diseases. In this study, 29 AMPs of varying length and characteristics were synthesised chemically and were evaluated for their ability to inhibit the growth of Bacillus globigii, Bacillus anthracis and Burkholderia thailandensis. Amongst the peptides tested, sheep myeloid antimicrobial peptide-29 (SMAP-29) was the most potent, inhibiting both B. globigii and B. anthracis at submicromolar concentrations. However, SMAP-29 was less effective against B. thailandensis (minimum inhibitory concentration of 71 microM). Haemolytic activity and cytotoxicity were determined using human blood cells and human embryonic kidney 293S cells, respectively. Most of the peptides tested showed varying degrees of haemolytic activity and cytotoxicity, with SMAP-29 being highly haemolytic and cytotoxic under the conditions tested. Nevertheless, strategies to reduce toxicity whilst maintaining high antimicrobial activity are worth pursuing in light of the results obtained.

Report of selective cortical infarcts in the primate clot model of vasospasm after subarachnoid hemorrhage.

BACKGROUND: In human autopsy studies, 70% to 80% of patients with aneurysmal subarachnoid hemorrhage (SAH) showed infarcts in cerebral cortex covered by subarachnoid blood. Thus far, no animal model of SAH is known to produce this peculiar infarct pattern, and its pathogenesis remains enigmatic. OBJECTIVE: To investigate whether such infarcts occur in the clot model of SAH in primates. METHODS: We performed a retrospective pathological review of 16 primate brains. In 13 cynomolgus monkeys, a blood clot was placed around the middle cerebral artery after additional removal of the arachnoid membrane from the basal surface of the frontal and temporal cortexes. Three animals underwent sham surgery without placement of a blood clot (controls). The brains were harvested between days 1 and 28 after SAH and examined by a neuropathologist blinded to study group. RESULTS: We identified 2 types of cortical infarcts. A band of selective cortical laminar necrosis parallel to the cortical surface ("horizontal") was found in 5 animals. The second category of cortical lesions had a "vertical" extension. It included wedge-shaped (n = 2) or pillarlike (n = 2) necrosis. Both horizontal and vertical infarcts were located exclusively in areas adjacent to subarachnoid blood. The presence of a cortical infarct did not correlate with the degree of middle cerebral artery vasospasm (r2 = .24, P = .13). CONCLUSION: The presence of cortical infarcts suggests that a modified nonhuman primate model of SAH is suitable to examine the pathogenesis of proximal vasospasm and permits investigation of cortical lesions similar to those reported in patients after SAH. Furthermore, it indicates that direct effects of the blood clot on the brain and microcirculation contribute to the development of cortical infarcts after SAH.

Effects of arterial and venous wall homogenates, arterial and venous blood, and different combinations to the cerebral vasospasm in an experimental model.

BACKGROUND: Risks related to rebleeding of a ruptured intracranial aneurysm have decreased. However, ischemic neurologic deficits related to vasospasm are still the leading causes of mortality and morbidity. It is well known that vasospasm is a dynamic process affected by various factors. The severity of vasospasm in animal models and clinical observations differ from each other. This variability has not been completely explained by blood and blood degradation products. Therefore, metabolites released from the damaged vessel wall during the bleeding are thought to play an important role in vasospasm. METHOD: To test this hypothesis, we used 46 male Wistar rats that were divided into 7 groups and administered one of the following to cisterna magna: venous blood, arterial blood, arterial wall homogenate, venous wall homogenate, combined mixture of arterial blood and artery wall homogenate, or combined mixture of venous blood and venous wall homogenate. Brainstems of the rats were excised, and the basilar arteries were harvested for morphometric measurements. RESULT: There were significant differences between the degree of vasospasm caused by arterial and venous blood (P < .0001). The intraluminal area of the basilar artery was significantly narrower after application of arterial blood, artery wall homogenate, or their combination (49% +/- 1%) than after venous groups (30% +/- 1.9%) (P < .0001). CONCLUSION: The results of this experiment demonstrated that metabolites from vessel walls play as important roles in the pathophysiology of vasospasm as blood and blood degradation products. Further investigation of these metabolites will improve our understanding of vasospasm, pathophysiology, and its treatment.

Protective effect of different parts of Cassia fistula on human umbilical vein endothelial cells against glycated protein-induced toxicity in vitro.

The protective effect of methanol extracts of Cassia fistula (flowers, leaves and bark) was examined in vitro in human umbilical vein endothelial cells (HUVEC) against toxicity induced by glycated protein (GFBS) in vitro. The experiments consisted of eight groups of HUVEC with five flasks in each group. Group I was treated with 15% FBS, group II with GFBS (70 microM) alone, and the other six groups were treated with GFBS plus 25 and 50 microg of each of the three types of C. fistula extracts. After 72 h of incubation, cells were collected and tested for lipid peroxidation, antioxidant enzyme activities and glutathione S-transferase (GST). The protective effect of C. fistula extracts against GFBS-induced cytotoxicity was examined in HUVEC by using trypan blue exclusion and MTT assays. Results showed that HUVEC incubated with GFBS alone showed a significant (P < 0.001) elevation of lipid peroxidation accompanied by depletion of superoxide dismutase (SOD), catalase, glutathione peroxidase (GPx) and glutathione reductase (GR), in addition to decreased cytosolic GST. Treatment of HUVEC with C. fistula extracts at a concentration of 25 and 50 microg significantly decreased lipid peroxidation and normalized the activities of the antioxidant enzymes and GST levels in a concentration-dependent manner. Morphological changes of HUVEC were compared with respective controls; in addition, the C. fistula extracts increased the viability of HUVEC damaged by GFBS. A protective effect of C. fistula extracts on HUVEC against GFBS-induced toxicity suggested a potential beneficial effect of the extract in preventing diabetic angiopathies.

SMAP29 congeners demonstrate activity against oral bacteria and reduced toxicity against oral keratinocytes.

INTRODUCTION: Cathelicidins are antimicrobial peptides found in epithelial and mucosal tissues as well as the secondary granules of neutrophils. SMAP29, a sheep cathelicidin, has differential antimicrobial properties against various pathogens, including periodontal organisms. The purpose of this study was to evaluate the antimicrobial properties and cytotoxicity of SMAP29, SMAP28, and three congeners (SMAP18A, SMAP18D, and SMAP14A). METHODS: The peptides at concentrations ranging from 0.25 to 250 microg/ml were tested for their activity against multiple strains of Streptococcus mutans, Streptococcus sanguis, Actinomyces israelii, Actinomyces naeslundii, Actinobacillus actinomycetemcomitans, Fusobacterium nucleatum, Peptostreptococcus micros, and Porphyromonas gingivalis using a radial diffusion assay. Cytotoxicity of keratinocytes was evaluated by measuring lactate dehydrogenase release after incubation with the individual peptides. RESULTS: SMAP28, thought to be the biologically active peptide, was the most potent antimicrobial (range of minimum inhibitory concentrations 0.06-7.03 microg/ml, P < 0.05); however, the activity of SMAP28 and SMAP29 was strongly associated (r = 0.933). The congeners also demonstrated antimicrobial activity against the bacteria tested (range of minimum inhibitory concnetrations 0.21-79 microg/ml). Overall, F. nucleatum was the most susceptible organism, while P. gingivalis was the least susceptible. Keratinocyte cytotoxicity was dependent on peptide length and dose. SMAP28 was the most cytotoxic, while SMAP14A was the least cytotoxic. CONCLUSION: The antimicrobial activities against oral microorganisms and the minimal toxicity seen in this study suggest that the congeners of SMAP29 may serve as an alternative to traditional antibiotics in the prevention and treatment of periodontal and other oral diseases.

Antimicrobial peptides versus invasive infections.

It has been estimated that there are more microorganisms within and upon the human body than there are human cells. By necessity, every accessible niche must be defended by innate mechanisms to prevent invasive infection, and ideally that precludes the need for robust inflammatory responses. Yet the potential for pathogens to transcend the integument actively or passively and access the bloodstream emphasizes the need for rapid and potent antimicrobial defense mechanisms within the vascular compartment. Antimicrobial peptides from leukocytes have long been contemplated as being integral to defense against these infections. Recently, platelets are increasingly recognized for their likely multiple roles in antimicrobial host defense. Platelets and leukocytes share many structural and functional archetypes. Once activated, both cell types respond in specific ways that emphasize key roles for their antimicrobial peptides in host defense efficacy: (a) targeted accumulation at sites of tissue injury or infection; (b) direct interaction with pathogens; and (c) deployment of intracellular (leukocyte phagosomes) or extracellular (platelet secretion) antimicrobial peptides. Antimicrobial peptides from these cells exert rapid, potent, and direct antimicrobial effects against organisms that commonly access the bloodstream. Experimental models in vitro and in vivo show that antimicrobial peptides from these cells significantly contribute to prevent or limit infection. Moreover, certain platelet antimicrobial proteins are multifunctional kinocidins (microbicidal chemokines) that recruit leukocytes to sites of infection, and potentiate the antimicrobial mechanisms of these cells. In turn, pathogens pre-decorated by kinocidins may be more efficiently phagocytosed and killed by leukocytes and their antimicrobial peptide arsenal. Hence, multiple and relevant interactions between platelets and leukocytes have immunologic functions yet to be fully understood. A clearer definition of these interactions, and the antimicrobial peptide effectors contributing to these functions, will significantly advance our understanding of antimicrobial host defense against invasive infection. In addition, this knowledge may accelerate development of novel anti-infective agents and strategies against pathogens that have become refractory to conventional antimicrobials.

Characterization of Bothrops jararaca coagulation inhibitor (BjI) and presence of similar protein in plasma of other animals.

BjI, a protein isolated from Bothrops jararaca snake blood, inhibits the coagulant activity of thrombin. This protein presents two bands of 109 and 138 kDa by SDS-PAGE under reducing conditions. In order to verify the presence of BjI-like proteins in plasma of other animals (reptiles and non-reptiles), we raised a specific polyclonal antibody in mice to it, and we verified immunological cross-reaction by western blotting, considering as positive reactions the development of bands with either 109 or 138 kDa. Similar proteins were identified in Bothrops neuwiedi and Crotalus durissus terrificus snakes. In contrast, no BjI-like protein in other classes of animals was noticeable, nor in other snakes tested. Interestingly, a prolonged thrombin time was found only in snake plasmas that showed similar BjI proteins. BjI bound to two proteins of B. jararaca venom, identified by western blotting. The N-terminal of the B. jararaca venom proteins showed similarity with thrombin-like proteins isolated from other snake venoms. In conclusion, there are similar proteins to BjI in plasmas of B. neuwiedi and Crotalus durissus terrificus, and these proteins also prolong thrombin time. Moreover, these results evidence the presence of target enzymes in snake venom for plasma BjI.

Signal peptide of eosinophil cationic protein is toxic to cells lacking signal peptide peptidase.

Eosinophil cationic protein (ECP) is a toxin secreted by activated human eosinophils. The properties of mature ECP have been well studied but those of the signal peptide of ECP (ECPsp) are not clear. In this study, several chimeric proteins containing N-terminal fusion of ECPsp were generated, and introduced into Escherichia coli, Pichia pastoris, and human epidermoid carcinoma cell line A431 to study the function of ECPsp. We found that expression of ECPsp chimeric proteins inhibited the growth of E. coli and P. pastoris but not A431 cells. Primary sequence analysis and in vitro transcription/translation of ECPsp have revealed that it is a potential substrate for human signal peptide peptidase (hSPP), an intramembrane protease located in endoplasmic reticulum. In addition, knockdown of the hSPP mRNA expression in ECPsp-eGFP/A431 cells caused the growth inhibitory effect, whereas complementally expression of hSPP in P. pastoris system rescued the cell growth. Taken together, we have demonstrated that ECPsp is a toxic signal peptide, and expression of hSPP protects the cells from growth inhibition.

rBPI(10-193) is secreted by CHO cells and retains the activity of rBPI21.

rBPI23, a recombinant N-terminal fragment of human bactericidal/permeability-increasing protein (BPI), kills Gram-negative bacteria and neutralizes endotoxin. rBPI21, a variant in which cysteine 132 is changed to alanine, retains the activities of rBPI23. Analysis of certain purified rBPI21 preparations revealed that some of the molecules had lost nine amino acids from the amino terminus. To explore the effect of this modification on structure and activity, we cloned and expressed a variant of rBPI21, designated rBPI(10-193), which lacks the first nine amino acids. A monoclonal antibody believed to recognize the amino terminus of rBPI21 cross-reacted with rBPI21, but not with rBPI(10-193) or full length recombinant BPI (rBPI). These results demonstrated that the antibody recognizes the first nine amino acids of rBPI21 and that this region of the holoprotein (rBPI) is inaccessible to the antibody (as suggested by the known 3-D structure). Purified rBPI(10-193) and rBPI21 were similarly potent in in vitro assays measuring bactericidal, endotoxin binding and neutralization activities. In a mouse model of lethal bacteremia, rBPI(10-193) and rBPI21 were similarly potent whereas in a mouse endotoxin challenge model, rBPI(10-193) appeared to be at least 2-fold more potent than rBPI21. In conscious rats, a rapid bolus dose of 40 mg/kg of rBPI21 caused a significant transient decrease in blood pressure while the same dose of rBPI(10-193) caused no blood pressure decrease. We conclude from these studies that the first nine amino acids of rBPI21 are not essential for the anti-infective and endotoxin-neutralizing activities of BPI.