Organotypic model of the gingiva for studying bacterial and viral pathogens implicated in periodontitis.

Background: Three-dimensional (3D) tissue models bridge the gap between conventional two-dimensional cell cultures and animal models. The aim of this study was to develop an organotypic 3D gingival (OTG) model to provide a tool to investigate bacterial and viral pathogens in periodontitis. Methods: The OTG model composed of gingival fibroblasts (GFs) and telomerase-immortalized gingival keratinocytes (TIGKs) was constructed and applied to study infections by Porphyromonas gingivalis and herpes simplex virus 1 (HSV-1). Immunohistochemical staining, confocal microscopy, qPCR, titration techniques, and colony-forming unit counts were applied to interrogate epithelial markers expression, monitor P. gingivalis and HSV-1 presence, and evaluate the immune response along with the efficiency of antimicrobial drugs. Results: The OTG model resembled the morphology of the human gingiva. During infection, both pathogens penetrated deep into the tissue and persisted for a few days with P. gingivalis also forming a biofilm on the cell surface. The infection triggered the expression of inflammatory mediators in cells and both pathogens were efficiently eliminated by specific antimicrobials. Conclusions: Presented OTG model constitutes a simple and convenient tool to study the interaction between bacterial and viral pathogens within the gingival tissue, including penetration, persistence and biofilm formation. It is also suitable to examine the efficiency of antimicrobial drugs.

Electrochemical detection of bacterial endotoxin lipopolysaccharide (LPS) on gold electrode modified with DAL-PEG-DK5-PEG-OH - Antimicrobial peptide conjugate.

This work describes fabrication of gold electrodes modified with peptide conjugate DAL-PEG-DK5-PEG-OH that enables ultra-sensitive detection of lipopolysaccharide (LPS) isolated from the reference strain of Escherichia coli O26:B6. The initial step of the established procedure implies immobilization of the fully protected DAL-PEG-DK5-PEG-OH peptide on the surface of the gold electrode previously modified by cysteamine. Then side chain- and Fmoc-deprotection was performed in situ on the electrode surface, followed by its incubation in 1 % of BSA solution to block non-specific bindings sites before LPS detection. The efficiency of the modification was confirmed by X-ray Photoelectron Spectroscopy (XPS) measurements. Additionally, the cyclic voltammetry (CV) and electrochemical impendance spectroscopy (EIS) were employed to monitor the effectiveness of each step of the modification. The obtained results confirmed that the presence of the surface-attached covalently bound peptide DAL-PEG-DK5-PEG-OH enables LPS detection by means of CV technique within the range from 5 × 10-13 to 5 × 10-4 g/mL in PBS solution. The established limit of detection (LOD) for EIS measurements was 4.93 × 10-21 g/mL with wide linear detection range from 5 × 10-21 to 5 × 10-14 g/mL in PBS solution. Furthermore, we confirmed the ability of the electrode to detect LPS in a complex biological samples, like mouse urine and human serum. The effectiveness of the electrodes in identifying LPS in both urine and serum matrices was confirmed for samples containing LPS at both 2.5 × 10-15 g/mL and 2.5 × 10-9 g/mL.

Screening and characterization of aptamers recognizing the periodontal pathogen Tannerella forsythia.

Periodontal disease is one of the most common forms of inflammation. It is currently diagnosed by observing symptoms such as gingival bleeding and attachment loss. However, the detection of biomarkers that precede such symptoms would allow earlier diagnosis and prevention. Aptamers are short oligonucleotides or peptides that fold into three-dimensional conformations conferring the ability to bind molecular targets with high affinity and specificity. Here we report the selection of aptamers that bind specifically to the bacterium Tannerella forsythia, a pathogen frequently associated with periodontal disease. Two aptamers with the highest affinity were examined in more detail, revealing that their binding is probably dependent on mirolysin, a surface-associated protease secreted by the T. forsythia type-9 secretion system. The aptamers showed minimal cross-reactivity to other periodontopathogens and are therefore promising leads for the development of new tools to study the composition of the periodontitis-associated dysbiotic bacteriome as well as inexpensive new diagnostic assays.

Identification and characterization of aptameric inhibitors of human neutrophil elastase.

Human neutrophil elastase (HNE) plays a pivotal role in innate immunity, inflammation, and tissue remodeling. Aberrant proteolytic activity of HNE contributes to organ destruction in various chronic inflammatory diseases including emphysema, asthma, and cystic fibrosis. Therefore, elastase inhibitors could alleviate the progression of these disorders. Here, we used the systematic evolution of ligands by exponential enrichment to develop ssDNA aptamers that specifically target HNE. We determined the specificity of the designed inhibitors and their inhibitory efficacy against HNE using biochemical and in vitro methods, including an assay of neutrophil activity. Our aptamers inhibit the elastinolytic activity of HNE with nanomolar potency and are highly specific for HNE and do not target other tested human proteases. As such, this study provides lead compounds suitable for the evaluation of their tissue-protective potential in animal models.

MCP-Induced Protein 1 Participates in Macrophage-Dependent Endotoxin Tolerance.

Endotoxin tolerance is a state of hyporesponsiveness to LPS, triggered by previous exposure to endotoxin. Such an immunosuppressive state enhances the risks of secondary infection and has been associated with the pathophysiology of sepsis. Although this phenomenon has been extensively studied, its molecular mechanism is not fully explained. Among candidates that play a crucial role in this process are negative regulators of TLR signaling, but the contribution of MCP-induced protein 1 (MCPIP1; Regnase-1) has not been studied yet. To examine whether macrophage expression of MCPIP1 participates in endotoxin tolerance, we used both murine and human primary macrophages devoid of MCPIP1 expression. In our study, we demonstrated that MCPIP1 contributes to LPS hyporesponsiveness induced by subsequent LPS stimulation and macrophage reprogramming. We proved that this mechanism revolves around the deubiquitinase activity of MCPIP1, which inhibits the phosphorylation of MAPK and NF-κB activation. Moreover, we showed that MCPIP1 controlled the level of proinflammatory transcripts in LPS-tolerized cells independently of its RNase activity. Finally, we confirmed these findings applying an in vivo endotoxin tolerance model in wild-type and myeloid MCPIP1-deficient mice. Taken together, this study describes for the first time, to our knowledge, that myeloid MCPIP1 participates in endotoxin tolerance and broadens the scope of known negative regulators of the TLR4 pathway crucial in this phenomenon.

Imaging of Clear Cell Renal Carcinoma with Immune Checkpoint Targeting Aptamer-Based Probe.

Immune checkpoint targeting immunotherapy has revolutionized the treatment of certain cancers in the recent years. Determination of the status of immune checkpoint expression in particular cancers may assist decision making. Here, we describe the development of a single-stranded aptamer-based molecular probe specifically recognizing human PD-L1. Target engaging aptamers are selected by iterative enrichment from a random ssDNA pool and the binding is characterized biochemically. Specificity and dose dependence is demonstrated in vitro in the cell culture using human kidney tumor cells (786-0), human melanoma cells (WM115 and WM266.4) and human glioblastoma LN18 cancer cells. The utility of the probe in vivo is demonstrated using two mouse tumor models, where we show that the probe exhibits excellent potential in imaging. We postulate that further development of the probe may allow universal imaging of different types of tumors depending on their PD-L1 status, which may find utility in cancer diagnosis.

The influence of OSPE and PBL on competency-based pharmacy student self-assessment.

BACKGROUND: In addition to vocational education and experience, the pharmaceutical profession's proper pursuit requires acquiring and continuously improving professional competencies. In recent years, the need has been increasingly highlighted for developing a medical education system based on helping students develop their competencies. It is necessary to adapt the tools and methods for assessing competencies during formal education. It will enable students to know the directions of further personal or professional development. OBJECTIVE: The study aimed to compare pharmacy students' self-assessment outcomes before and after the Objective Structured Practical Examination (OSPE), which finished the Pharmaceutical Care course (PCc). The study's purpose was also to compare the outcomes of the self-assessment of competencies between the students of two academic years for whom classes on the PCc were provided by different methods. METHODS: The study was conducted over two academic years (2018/2019 and 2019/2020) among 5th-year students enrolled at the Faculty of Pharmacy of the Jagiellonian University Medical College (JUMC) at the end of the Pharmaceutical Care course. Different teaching methods were used in the delivery of the course in these academic years. The students self-assessed their competencies using a questionnaire consisting of a list of personal and patient care competencies. The students completed the questionnaire before and after the OSPE, which followed the completion of the PCc. RESULTS: Students' professional competencies as self-assessed after the exam were higher than those assessed before the exam. Differences were observed in both personal and patient care competencies. Students taking the course in the 2019/2020 academic year set their pre-OSPE competencies higher than students taking the PCc in 2018/2019. CONCLUSION: The self-assessment scores increased for most competencies included in the study following the OSPE. This may suggest that taking part in the exam, involvement in patient's case simulations, and self-assessment of performance at individual stages of the exam contributed to increased subjective assessment of professional competencies.

Lipidation of Temporin-1CEb Derivatives as a Tool for Activity Improvement, Pros and Cons of the Approach.

The alarming raise of multi-drug resistance among human microbial pathogens makes the development of novel therapeutics a priority task. In contrast to conventional antibiotics, antimicrobial peptides (AMPs), besides evoking a broad spectrum of activity against microorganisms, could offer additional benefits, such as the ability to neutralize toxins, modulate inflammatory response, eradicate bacterial and fungal biofilms or prevent their development. The latter properties are of special interest, as most antibiotics available on the market have limited ability to diffuse through rigid structures of biofilms. Lipidation of AMPs is considered as an effective approach for enhancement of their antimicrobial potential and in vivo stability; however, it could also have undesired impact on selectivity, solubility or the aggregation state of the modified peptides. In the present work, we describe the results of structural modifications of compounds designed based on cationic antimicrobial peptides DK5 and CAR-PEG-DK5, derivatized at their N-terminal part with fatty acids with different lengths of carbon chain. The proposed modifications substantially improved antimicrobial properties of the final compounds and their effectiveness in inhibition of biofilm development as well as eradication of pre-formed 24 h old biofilms of Candida albicans and Staphylococcus aureus. The most active compounds (C5-DK5, C12-DK5 and C12-CAR-PEG-DK5) were also potent against multi-drug resistant Staphylococcus aureus USA300 strain and clinical isolates of Pseudomonas aeruginosa. Both experimental and in silico methods revealed strong correlation between the length of fatty acid attached to the peptides and their final membranolytic properties, tendency to self-assemble and cytotoxicity.

Citrullination-Resistant LL-37 Is a Potent Antimicrobial Agent in the Inflammatory Environment High in Arginine Deiminase Activity.

LL-37, the only member of the mammalian cathelicidin in humans, plays an essential role in innate immunity by killing pathogens and regulating the inflammatory response. However, at an inflammatory focus, arginine residues in LL-37 can be converted to citrulline via a reaction catalyzed by peptidyl-arginine deiminases (PAD2 and PAD4), which are expressed in neutrophils and are highly active during the formation of neutrophil extracellular traps (NETs). Citrullination impairs the bactericidal activity of LL-37 and abrogates its immunomodulatory functions. Therefore, we hypothesized that citrullination-resistant LL-37 variants would retain the functionality of the native peptide in the presence of PADs. To test this hypothesis, we synthetized LL-37 in which arginine residues were substituted by homoarginine (hArg-LL-37). Bactericidal activity of hArg-LL-37 was comparable with that of native LL-37, but neither treatment with PAD4 nor exposure to NETs affected the antibacterial and immunomodulatory activities of hArg-LL-37. Importantly, the susceptibilities of LL-37 and hArg-LL-37 to degradation by proteases did not significantly differ. Collectively, we demonstrated that citrullination-resistant hArg-LL-37 is an attractive lead compound for the generation of new agents to treat bacterial infections and other inflammatory diseases associated with enhanced PAD activity. Moreover, our results provide a proof-of-concept for synthesis of therapeutic peptides using homoarginine.

The Potential Role of the Pharmacist in Supporting Patients with Depression - A Literature-Based Point of View.

OBJECTIVE: The current article is aimed at identifying the best practice for counseling around depression in community and outpatient pharmacies, resulting in a draft guideline, proposing key steps and an algorithm for integration of community pharmacists into care for patients with depression. METHODS: A literature review was performed followed by a detailed analysis, for the purpose of creation a short draft document used as a basis for creation of a guideline for pharmaceutical care for patients with depression. The technological scheme PRISMA flow diagram was applied. The paper is based on current knowledge, taking into consideration already published articles, guidelines, and recommendations about pharmaceutical care for patients with depression, giving a basis for further studies. RESULTS: This paper includes two main sections: 1) depression - a short description of the main symptoms, risk factors and pharmacotherapy guidelines available in Bulgaria important for the purposes of ensuring qualitative community-based pharmaceutical care; and 2) the pharmacists' role in providing high-quality care - the main aspects of pharmaceutical care for patients with depression with specific examples. CONCLUSION: The involvement of pharmacists in supporting depressive patients is crucial taking into account the specific characteristics of the pharmacological treatment: delayed onset of clinical results, risks in case of sudden pharmacotherapy abruption without physician consultation, multiple adverse drug reactions and drug-drug, drug-food and drug-alcohol interactions, etc. The current article could also be used as an initial document for creating a methodological guideline for providing pharmaceutical care services for patients with depression.

The Bactericidal Activity of Temporin Analogues Against Methicillin Resistant Staphylococcus aureus.

Staphylococcus aureus is a major infectious agent responsible for a plethora of superficial skin infections and systemic diseases, including endocarditis and septic arthritis. Recent epidemiological data revealed the emergence of resistance to commonly used antibiotics, including increased numbers of both hospital- and community-acquired methicillin-resistant S. aureus (MRSA). Due to their potent antimicrobial functions, low potential to develop resistance, and immunogenicity, antimicrobial peptides (AMPs) are a promising alternative treatment for multidrug-resistant strains. Here, we examined the activity of a lysine-rich derivative of amphibian temporin-1CEb (DK5) conjugated to peptides that exert pro-proliferative and/or cytoprotective activity. Analysis of a library of synthetic peptides to identify those with antibacterial potential revealed that the most potent agent against multidrug-resistant S. aureus was a conjugate of a temporin analogue with the synthetic Leu-enkephalin analogue dalargin (DAL). DAL-PEG-DK5 exerted direct bactericidal effects via bacterial membrane disruption, leading to eradication of both planktonic and biofilm-associated staphylococci. Finally, we showed that accumulation of the peptide in the cytoplasm of human keratinocytes led to a marked clearance of intracellular MRSA, resulting in cytoprotection against invading bacteria. Collectively, the data showed that DAL-PEG-DK5 might be a potent antimicrobial agent for treatment of staphylococcal skin infections.

Effect of Education for Hypertensive Patients with Correctly Performed Self-Blood Pressure Monitoring (SBPM).

INTRODUCTION: The study objective was to assess the impact of pharmacist-led education on the patient's knowledge and skills on SBPM (self-blood pressure monitoring). METHODS: Patient knowledge was assessed using tests and patients' skills were based on a checklist (20 parameters) completed by the researcher based on the SBPM video records. Patients taking pressure measurements were filmed for 20 days and after 6 months. After the first 10 days, patients were educated about the correct SBPM procedure. Knowledge tests were repeated three times (before/after/6 months after education). RESULTS: All patients' knowledge and skills in the field of SBPM were improved after education. After the education, patients scored an average of 9 out of 10 points in the knowledge test (increasing an average of five points after education), six months later-an average increase of 7.36 points. Patient skills after training were rated at 17.4 out of 20 points on average (increase by an average of 5.14 points after education), six months later, there was an average of 16.23 points. CONCLUSIONS: The study showed an increase in patients' knowledge and skills in the field of SBPM after training.

Are We Ready to Implement Competence-Based Teaching in Pharmacy Education in Poland?

Pharmacists in Poland are responsible for the dispensing and quality control of pharmaceuticals. The education process in pharmacy is regulated and monitored at the national level. Pharmacy education at Jagiellonian University is organized in a traditional way based on input and content teaching. The aim of the study was to determinate whether the Jagiellonian University curriculum in the Pharmacy program meets the criteria of the European Competence Framework. The mapping of the intended curriculum was done by four academic teachers. The qualitative and quantitative analysis of the distribution of the European Competence Framework among a group of courses and study years was done. We observed that most of the personal competencies are offered to students in their senior years, while the patient care competencies are distributed equally during the cycle of the study, and only some of them are overrepresented at the senior years. We need a legislation change at the national level as well as organizational and mental change at the university level to move from learning outcome-based pharmacy education to competence-based.

Novel Polyanions Inhibiting Replication of Influenza Viruses.

Novel sulfonated derivatives of poly(allylamine hydrochloride) (NSPAHs) and N-sulfonated chitosan (NSCH) have been synthesized, and their activity against influenza A and B viruses has been studied and compared with that of a series of carrageenans, marine polysaccharides of well-documented anti-influenza activity. NSPAHs were found to be nontoxic and very soluble in water, in contrast to gel-forming and thus generally poorly soluble carrageenans.In vitroandex vivostudies using susceptible cells (Madin-Darby canine kidney epithelial cells and fully differentiated human airway epithelial cultures) demonstrated the antiviral effectiveness of NSPAHs. The activity of NSPAHs was proportional to the molecular mass of the chain and the degree of substitution of amino groups with sulfonate groups. Mechanistic studies showed that the NSPAHs and carrageenans inhibit influenza A and B virus assembly in the cell.

Human Coronavirus HKU1 Spike Protein Uses O-Acetylated Sialic Acid as an Attachment Receptor Determinant and Employs Hemagglutinin-Esterase Protein as a Receptor-Destroying Enzyme.

UNLABELLED: Human coronavirus (hCoV) HKU1 is one of six hCoVs identified to date and the only one with an unidentified cellular receptor. hCoV-HKU1 encodes a hemagglutinin-esterase (HE) protein that is unique to the group a betacoronaviruses (group 2a). The function of HKU1-HE remains largely undetermined. In this study, we examined binding of the S1 domain of hCoV-HKU1 spike to a panel of cells and found that the S1 could specifically bind on the cell surface of a human rhabdomyosarcoma cell line, RD. Pretreatment of RD cells with neuraminidase (NA) and trypsin greatly reduced the binding, suggesting that the binding was mediated by sialic acids on glycoproteins. However, unlike other group 2a CoVs, e.g., hCoV-OC43, for which 9-O-acetylated sialic acid (9-O-Ac-Sia) serves as a receptor determinant, HKU1-S1 bound with neither 9-O-Ac-Sia-containing glycoprotein(s) nor rat and mouse erythrocytes. Nonetheless, the HKU1-HE was similar to OC43-HE, also possessed sialate-O-acetylesterase activity, and acted as a receptor-destroying enzyme (RDE) capable of eliminating the binding of HKU1-S1 to RD cells, whereas the O-acetylesterase-inactive HKU1-HE mutant lost this capacity. Using primary human ciliated airway epithelial (HAE) cell cultures, the only in vitro replication model for hCoV-HKU1 infection, we confirmed that pretreatment of HAE cells with HE but not the enzymatically inactive mutant blocked hCoV-HKU1 infection. These results demonstrate that hCoV-HKU1 exploits O-Ac-Sia as a cellular attachment receptor determinant to initiate the infection of host cells and that its HE protein possesses the corresponding sialate-O-acetylesterase RDE activity. IMPORTANCE: Human coronaviruses (hCoV) are important human respiratory pathogens. Among the six hCoVs identified to date, only hCoV-HKU1 has no defined cellular receptor. It is also unclear whether hemagglutinin-esterase (HE) protein plays a role in viral entry. In this study, we found that, similarly to other members of the group 2a CoVs, sialic acid moieties on glycoproteins are critical receptor determinants for the hCoV-HKU1 infection. Interestingly, the virus seems to employ a type of sialic acid different from those employed by other group 2a CoVs. In addition, we determined that the HKU1-HE protein is an O-acetylesterase and acts as a receptor-destroying enzyme (RDE) for hCoV-HKU1. This is the first study to demonstrate that hCoV-HKU1 uses certain types of O-acetylated sialic acid residues on glycoproteins to initiate the infection of host cells and that the HKU1-HE protein possesses sialate-O-acetylesterase RDE activity.

The nucleocapsid protein of human coronavirus NL63.

Human coronavirus (HCoV) NL63 was first described in 2004 and is associated with respiratory tract disease of varying severity. At the genetic and structural level, HCoV-NL63 is similar to other members of the Coronavirinae subfamily, especially human coronavirus 229E (HCoV-229E). Detailed analysis, however, reveals several unique features of the pathogen. The coronaviral nucleocapsid protein is abundantly present in infected cells. It is a multi-domain, multi-functional protein important for viral replication and a number of cellular processes. The aim of the present study was to characterize the HCoV-NL63 nucleocapsid protein. Biochemical analyses revealed that the protein shares characteristics with homologous proteins encoded in other coronaviral genomes, with the N-terminal domain responsible for nucleic acid binding and the C-terminal domain involved in protein oligomerization. Surprisingly, analysis of the subcellular localization of the N protein of HCoV-NL63 revealed that, differently than homologous proteins from other coronaviral species except for SARS-CoV, it is not present in the nucleus of infected or transfected cells. Furthermore, no significant alteration in cell cycle progression in cells expressing the protein was observed. This is in stark contrast with results obtained for other coronaviruses, except for the SARS-CoV.

Porphyromonas gingivalis enzymes enhance infection with human metapneumovirus in vitro.

Relatively recently discovered, human metapneumovirus (HMPV) is a human pathogen with worldwide prevalence, accounting for a substantial percentage of respiratory tract diseases. Concurrent viral and bacterial infections enable intricate mechanisms of cooperation between pathogens, which complicate the symptoms and outcome of the disease. Such bilateral interactions are based on the modulation of bacterial growth on epithelium pathologically altered during viral illness and the modulation of immune responses, as well as the enhancement of virus replication by bacterial virulence factors. This study showed that proteases produced by Porphyromonas gingivalis, a Gram-negative bacterium implicated in the development of periodontitis, named gingipains, facilitated HMPV replication in LLC-MK2 cells and may contribute to HMPV pathogenicity in patients with periodontitis. Gingipains at low nanomolar concentrations enabled HMPV replication and allowed virus propagation in vitro. In contrast to previously published data for influenza virus, however, Staphylococcus aureus proteases and human neutrophil elastase did not affect virus replication.

Infection with human coronavirus NL63 enhances streptococcal adherence to epithelial cells.

Understanding the mechanisms of augmented bacterial pathogenicity in post-viral infections is the first step in the development of an effective therapy. This study assessed the effect of human coronavirus NL63 (HCoV-NL63) on the adherence of bacterial pathogens associated with respiratory tract illnesses. It was shown that HCoV-NL63 infection resulted in an increased adherence of Streptococcus pneumoniae to virus-infected cell lines and fully differentiated primary human airway epithelium cultures. The enhanced binding of bacteria correlated with an increased expression level of the platelet-activating factor receptor (PAF-R), but detailed evaluation of the bacterium-PAF-R interaction revealed a limited relevance of this process.

Myeloperoxidase-catalyzed oxidative inactivation of human kininogens: the impairment of kinin-precursor and prekallikrein-binding functions.

Bradykinin-related vasoactive peptides (kinins) are important mediators of local and systemic inflammatory reactions. However, at local inflammatory foci, the production of kinins from proteinaceous precursors (kininogens) can be affected by reactive oxygen species released by phagocyte cells. One of the predominant oxidants at these places is hypochlorous acid which is formed from hydrogen peroxide and chloride ions by neutrophil myeloperoxidase. In this study, inactivation of human kininogens after oxidation with the myeloperoxidase-H2O2-chloride system was observed and analyzed by protein chemistry methods. The kinin release from oxidized kininogens by major kinin-producing enzymes, plasma and tissue kallikreins, proceed with a very low rate. This effect was assigned to apparent inability of kallikreins to process the kinin N-terminus owing to the conversion of the adjacent Met-361 residue to methionine sulfoxide. Additionally, the oxidized high-molecular mass kininogen lost its natural ability to bind plasma prekallikrein. This effect was assigned to the oxidation of Trp-569 residue within the prekallikrein-binding region which is subsequently destructed owing to cleavage of the peptide bond after that residue. One possible pathophysiological consequence of the described effects on kininogens could be the impairment of the normal assembly and triggering of the kinin-forming system on defense cell surfaces.

Degradation of human kininogens with the release of kinin peptides by extracellular proteinases of Candida spp.

The secretion of proteolytic enzymes by pathogenic microorganisms is one of the most successful strategies used by pathogens to colonize and infect the host organism. The extracellular microbial proteinases can seriously deregulate the homeostatic proteolytic cascades of the host, including the kinin-forming system, repeatedly reported to be activated during bacterial infection. The current study assigns a kinin-releasing activity to secreted proteinases of Candida spp. yeasts, the major fungal pathogens of humans. Of several Candida species studied, C. parapsilosis and C. albicans in their invasive filamentous forms are shown to produce proteinases which most effectively degrade proteinaceous kinin precursors, the kininogens. These enzymes, classified as aspartyl proteinases, have the highest kininogen-degrading activity at low pH (approx. 3.5), but the associated production of bradykinin-related peptides from a small fraction of kininogen molecules is optimal at neutral pH (6.5). The peptides effectively interact with cellular B2-type kinin receptors. Moreover, kinin-related peptides capable of interacting with inflammation-induced B1-type receptors are also formed, but with a reversed pH dependence. The presented variability of the potential extracellular kinin production by secreted aspartyl proteinases of Candida spp. is consistent with the known adaptability of these opportunistic pathogens to different niches in the host organism.