The effect of aromatic side chains on the dilational rheological properties of N-acyltaurate amphiphiles at water-decane interfaces.

To elucidate the effect of aromatic side chains on dilational rheological properties of N-acyltaurate amphiphiles at the decane-water interface, the interfacial rheological properties of sodium N-2-(2-naphthoxy)-tetradecanoyltaurinate (12+N-T) and sodium N-2-(p-butylphenoxy)-tetradecanoyltaurinate (12+4B-T) were investigated utilizing the drop shape analysis method. The effects of adsorption time, interfacial pressure, oscillating frequency, and bulk concentration on the interfacial dilational modulus and phase angle were explored. The results show that the 12+4B-T molecule with a longer hydrophobic chain shows a higher ability for reducing the interfacial tension (IFT). In addition, the interfacial films of both 12+N-T and 12+4B-T are dominated by diffusion exchange at high concentrations. The rigidity of molecules controls the diffusion exchange at low concentrations, while the molecular hydrodynamic radius determines the diffusion exchange at high concentrations. Thus, at low concentrations, the stronger intermolecular interaction between 12+4B-T molecules results in higher dilational modulus values than 12+N-T. When approaching the CMC (critical micelle concentration) value, the rapid diffusion exchange of 12+4B-T between the sublayer micelles and the interface causes a significant decrease in the dilational modulus, while the relatively rigid structure of 12+N-T enables a higher dilational modulus than 12+4B-T. What's more, the longer hydrophobic chain allows 12+4B-T molecules to escape from the interface more easily, resulting in a higher phase angle at low concentrations. However, the diffusion exchange of 12+4B-T is slower than that of 12+N-T, which results in lower phase angles for 12+4B-T than 12+N-T at high concentrations. In general, the introduction of a rigid naphthalene ring in the molecular structure gives the interfacial film greater strength at high concentration. The research results in this paper provide a new technique for the strength regulation of interfacial surfactant adsorption films.

Adsorption behaviors of novel betaines on the wettability of the quartz surface.

The contact angle measurements for the aqueous solutions of two pairs of zwitterions on quartz surfaces have been investigated by the sessile drop analysis. The different physicochemical parameters such as the critical micelle concentration (CMC), surface tension, contact angle, surface excess on air-liquid and solid-liquid interfaces and work of adhesion have been estimated. The obtained results show that the contact angle of surfactants such as alkyl carboxylbetaine (ACB) and ditolyl substituted alkyl carboxylbetaine (BCB) remains almost constant in a wide range of surfactant concentration and increases gradually above CMC, which are quite different from traditional surfactants reported in the literature. Surfactants with bigger polar groups have a more steric effect on the quartz surface and the contact angle remains relatively unchanged. Moreover, an increase in quartz-liquid interfacial tension (γSL) has been observed due to the adsorption of four zwitterionic surfactants. Especially for ACB and BCB, at the surfactant concentrations higher than 5 × 10(-5) mol L(-1), a moderate increase in the interfacial tension of the quartz-liquid is observed, which suggests that ACB and BCB can form a saturated adsorption film briefly on the quartz surface and then adsorb again. However, the addition of alkyl sulfobetaine (ASB) and ditolyl substituted alkyl sulfobetaine (BSB) after CMC cannot adsorb on the quartz surface again due to the steric effect of bigger polar groups.

Anthropometric measurements in 126 microtia reconstructions.

BACKGROUND: Due to its complex, three-dimensional morphology, auricular reconstruction remains one of the most challenging procedures in reconstructive surgery. A subject that remains controversial, however, is the question of the growth potential of the cartilaginous framework. This study explored the anthropometric changes of the reconstructed auricle and the contralateral normal ear in a series of Asian patients. METHODS: The records of 126 unilateral microtia patients in three age groups who underwent autogenous costal cartilage auricular reconstruction between 2007 and 2010 were reviewed. The average age was 14 years, and the average follow-up was 2.5 years. The auricular height was measured as the distance from the supra-auricle to the subauricle, and the width was measured as the distance from preauricle to the postauricle. Measurements of the height and width of the reconstructed auricle and the contralateral normal side were taken at implantation and at the final follow-up. Comparisons between the three age groups were performed using a paired Student t test to examine the mean auricular heights and widths for significant interval changes in size. RESULTS: The measurements showed average width increases of 1.24 mm (3.68%) and 1.35 mm (3.91%) in the reconstructed auricles of children and adolescents, respectively, but we did not find obvious changes in the adult group. No significant differences were found in the height measurement of the reconstructed auricle in the three groups. The mature size of the normal ear was reached by age 12, with slowing as patients entered adolescence and adulthood. Comparison of the reconstructed auricle and the contralateral normal ear revealed a very similar growth rate in the adult group. There were average height decreases of 0.77 mm (1.3%) and 1.3 mm (2.09%) in the reconstructed auricles of children and adolescents compared with the normal side. The results showed an average width increase of 1.13 mm (3.15%) in the adolescent group but not in the child or adult groups. CONCLUSIONS: The results of this study have generated some useful parameters for the study of growth of the reconstructed auricle and contralateral normal ear. This information serves to clarify the issue of auricular framework fabrication in terms of proper size, especially for Asian patients. Further investigation and analysis are necessary to provide further proof of the graft change.

Adsorption behaviors of cationic surfactants and wettability in polytetrafluoroethylene-solution-air systems.

Measurements of the advancing contact angle (θ) and adsorption properties were carried out for aqueous solutions of four cationic surfactants, hexadecanol glycidyl ether ammonium chloride (C(16)PC), Guerbet alcohol hexadecyl glycidyl ether ammonium chloride (C(16)GPC), hexadecanol polyoxyethylene(3) glycidyl ether ammonium chloride(C(16)(EO)(3)PC), and Guerbet alcohol hexadecyl polyoxyethylene(3) glycidyl ether ammonium chloride (C(16)G(EO)(3)PC), on the polytetrafluoroethylene (PTFE) surface using the sessile drop analysis. The obtained results indicate that the contact angle decreases to a minimum with the increasing concentration for all cationic surfactants. Surfactants with branched chain show lower θ values. Moreover, an increase of adhensional tension on the PTFE-water interface has been observed for the four cationic surfactants, and the branched ones have larger increases of adhensional tension. It is very interesting that the sharp decrease of θ appears mainly after critical micelle concentration (cmc) for C(16)GPC, C(16)(EO)(3)PC, and C(16)G(EO)(3)PC, which is quite different from traditional cationic surfactants reported in the literature. Especially for C(16)G(EO)(3)PC, there are two saturated adsorption stages on PTFE surface after cmc (which means the saturated adsorption film at air-solution interface has been formed). In the first saturated stage, the C(16)G(EO)(3)PC molecules are oriented parallel to the PTFE surface with saturated monolayer formed through hydrophobic interaction and hydrogen bond. In the second saturated stage, the hemimicelle has been formed on the PTFE surface, which can be supported by the QCM-D and SPR measurements.

Dilational Rheological Properties of Surfactants at the Crude Oil-Water Interface: The Effect of Branch-Preformed Particle Gels and Polymers.

The interfacial properties of a heterogeneous composite flooding system containing a surfactant fatty alcohol polyoxyethylene carboxylate (C12EO3C), branched-preformed particle gel (B-PPG), and polymer partly hydrolyzed polyacrylamide (HPAM) at the crude oil-water interface were investigated by a dilational rheology method. The results demonstrated that the C12EO3C molecules can form an elastic interfacial film with certain strength at the crude oil-water interface. The addition of HPAM to the C12EO3C solution has a detrimental effect on the interfacial film formed by C12EO3C molecules, leading to a decrease in the dilational modulus and an increase in the phase angle. Moreover, the addition of B-PPG to the C12EO3C solution also disrupts the stability and strength of the interfacial film of C12EO3C. In particular, linear HPAM with a lower steric hindrance is more likely to insert into the interfacial film of C12EO3C; thus, HPAM possesses a stronger destruction ability for the interfacial film of C12EO3C than B-PPG. When HPAM is compounded with B-PPG, a superimposed effect exists to cause more severe disruption for the interfacial film. The heterogeneous composite flooding system not only enhances oil recovery by increasing the viscosity of the bulk phase but also weakens the interfacial film to facilitate the post-treatment of the recovered crude oil. Thus, the heterogeneous composite flooding system exhibits promising prospects in practical application.

Massive true thymic hyperplasia in a 3-month-old infant: case report and literature review.

BACKGROUND: True thymic hyperplasia (TTH) is characterized as a distinct increase in both size and weight of thymus, which retains normal microscopic and immunohistochemical appearances. Massive true thymic hyperplasia (MTTH) is an extremely rare but significant subtype of TTH in pediatric ages due to its potentially serious consequences. It was reported that the age of cases with MTTH was predominantly between 1 and 15 years, while those before 1 year rarely occurred. By presenting the diagnosis and treatment process of our case as well as reviewing the related literature, we aimed to analyze the clinical characteristics of MTTH for patients younger than 1 year. CASE: A 3-month-old male infant was admitted to our department with a chief complaint of gradually increasing polypnea over 9 days, whose preoperative imaging examination showed a large intrathoracic soft tissue shadow predominantly on the right side. The percutaneous fine-needle biopsy guided by ultrasonography was performed to identify its diagnosis. However, proliferating lymphocytes and Hassall`s corpuscles were seen microscopically in the biopsy tissues, which were immunohistochemically positive for CD3, CD19, CD20, CD99, TdT, PCK and Ki67 ( > 90%). Due to the aggravating symptoms, a second operation with total thymectomy was carried out successfully for this infant, which confirmed the diagnosis of TTH again by both morphological study and immunohistochemical staining from the surgical specimen. CONCLUSIONS: By reviewing the literature, there were only 10 cases with MTTH reported between 1975 and 2020 for children aged < 1 year of life, together with our present one. In MTTH patient`s sex had an obviously male predominance (70%). Nine out of 10 presented initial symptoms or signs related to respiratory system and 6 patients showed respiratory distress. All patients were successfully treated by surgical thymectomy without any postoperative complications. The prognosis of MTTH was very successful.

Wettability of a quartz surface in the presence of four cationic surfactants.

Advancing contact angle (θ) measurements were carried out for aqueous solutions of four cationic surfactants, hexadecanol glycidyl ether ammonium chloride (C(16)PC), guerbet alcohol hexadecyl glycidyl ether ammonium chloride (C(16)GPC), hexadecanol polyoxyethylene(3) glycidyl ether ammonium chloride (C(16)(EO)(3)PC), and guerbet alcohol hexadecyl polyoxyethylene(3) glycidyl ether ammonium chloride (C(16)G(EO)(3)PC), on the quartz surface using the sessile drop analysis. The influences of surfactant type and bulk concentration on contact angle were expounded, and the changes in adhesional tension and adhesion work were discussed. The contact angle increases up to a maximum with the increasing concentration for all cationic surfactants. Surfactants with branched chain have more hydrophobic group density on the quartz surface, which results in higher values of maxima in contact angle curves. When ethylene oxide groups CH(2)CH(2)O were incorporated in the hydrophobic group, the decrease in contact angle maximum was observed for C(16)(EO)(3)PC and C(16)G(EO)(3)PC. Moreover, an increase in quartz-water interfacial free energy (γ(SL)) has been observed due to the adsorption of four cationic surfactants. The four cationic surfactants can form a monolayer with alignment structure on the quartz surface through electrostatic interaction and then form the bilayer with increasing bulk concentration. In contrast with literature, the maximal contact angles may not necessarily correspond to the beginning of the formation of bilayer for cationic surfactants at the quartz-water interface. Moreover, the concentrations corresponding to maximal contact angles for C(16)PC and C(16)(EO)(3)PC were much lower than their CMC. The contact angle passes through a maximum at a concentration obviously higher than CMC for C(16)G(EO)(3)PC.

[Study on the interaction between ICT fluorescence probe and bovine serum albumins].

The present article studied the interaction between intramolecular charge transfer fluorescence probe-1-keto-2-(p-dimethylaminobenzal)-tetrohydronaphthalene (KDTN) and bovine serum albumins (BSA). With the concentration of KDTN increasing, the fluorescence of BSA rapidly quenched and the fluorescence peak gradually blue-shifted. The result indicated that they were bound mainly by hydrophobic interaction. The binding sites is 0.94 (3 degrees C) and the equilibrium constant K is 3.27 x 10(4) L x mol(-1). Temperature increment is advantageous to the combination. It is a single static quenching process that the fluorescence of BSA quenches, which is induced by the combination of KDTN and BSA. Further study showed that different substances had different effects on the combination of KDTN and BSA.

Humic substances act as electron acceptor and redox mediator for microbial dissimilatory azoreduction by Shewanella decolorationis S12.

The potential for humic substances to serve as terminal electron acceptors in microbial respiration and the effects of humic substances on microbial azoreduction were investigated. The dissimilatory azoreducing microorganism Shewanella decolorationis S12 was able to conserve energy to support growth from electron transport to humics coupled to the oxidation of various organic substances or H2. Batch experiments suggested that when the concentration of anthraquinone-2-sulfonate (AQS), a humics analog, was lower than 3 mmol/l, azoreduction of strain S12 was accelerated under anaerobic condition. However, there was obvious inhibition to azoreduction when the concentration of the AQS was higher than 5 mmol/l. Another humics analog, anthraquinone-2-sulfonate (AQDS), could still prominently accelerate azoreduction, even when the concentration was up to 12 mmol/l, but the rate of acceleration gradually decreased with the increasing concentration of the AQDS. Toxic experiments revealed that AQS can inhibit growth of strain S12 if the concentration past a critical one, but AQDS had no effect on the metabolism and growth of strain S12 although the concentration was up to 20 mmol/l. These results demonstrated that a low concentration of humic substances not only could serve as the terminal electron acceptors for conserving energy for growth, but also act as redox mediator shuttling electrons for the anaerobic azoreduction by S. decolorationis S12. However, a high concentration of humic substances could inhibit the bacterial azoreduction, resulting on the one hand from the toxic effect on cell metabolism and growth, and on the other hand from competion with azo dyes for electrons as electron acceptor.

Engineering of the human vessel wall with hair follicle stem cells in vitro.

Hair follicle stem cells (HFSCs) are increasingly used as a stem cell paradigm in vascular tissue engineering due to the fact that they are a rich source of easily accessible multipotent adult stem cells. Promising results have been demonstrated with small diameter (less than 6 mm) tissue engineered blood vessels under low blood pressure, however engineering large vessels (>6 mm in diameter) remains a challenge due to the fact it demands a higher number of seed cells and higher quality biomechanical properties. The aim of the current study was to engineer a large vessel (6 mm in diameter) with differentiated smooth muscle cells (SMCs) induced from human (h)HFSCs using transforming growth factor­ß1 and platelet­derived growth factor BB in combination with low­serum culture medium. The cells were seeded onto polyglycolic acid and then wrapped around a silicone tube and further cultured in vitro. A round vessel wall was formed subsequent to 8 weeks of culture. Histological examination indicated that layers of smooth muscle­like cells and collagenous fibres were oriented in the induced group. In contrast, disorganised cells and collagenous fibres were apparent in the undifferentiated group. The approach developed in the current study demonstrated potential for constructing large muscular vessels with differentiated SMCs induced from hHFSCs.

Identification, structural characterization and expression analysis of a novel carbonic anhydrase from freshwater mussel Hyriopsis cumingii.

In this study, an α-carbonic anhydrase (α-CA), HcCA3, from Hyriopsis cumingii was characterized. The full-length cDNA of HcCA3 was 1628bp, including a CA domain and an ORF of 1053bp which encoded 350 amino acids. Its predicted molecular weight was 39.69kDa and the pI was 5.92. qRT-PCR was used to determine the expression of the gene in various tissues at 0h, 3h, 6h, 12h, 24h, 48h, 96h, 7d, 14d, 21d, 28d and 35d after inserting the pearl nucleus. The results showed that the HcCA3 was highly expressed in the mantle, whereas its expression was low in other tissues. Expression in the posterior mantle pallial (pMP) was significantly higher than that in the anterior mantle pallial (aMP) and mantle center (MC). Expression in the aMP, pMP and MC was significantly higher in purple mussels compared with that in white mussels. At the same time, during the formation of pearls, expression in the aMP, pMP and pearl sac (PS) decreased and then increased; whereas expression in the MC increased and then decreased. In-situ hybridization showed that the HcCA3 was expressed in both inside and outside epidermal cells. In protein level, Western blot showed that HcCA3 was mainly expressed in the aMP, pMP and MC. Our results suggest that HcCA3 play a role in the formation of shell and pearl sac formation.

[The effects of the humic substances on Azoreduction by Shewanella spp].

Under anaerobic conditions, Shewanella cinicaD14(T), Shewanella baltica and Shewane-lla putrefaciens are capable of high-rate azoreduction and humus reduction. The results indicated that at low concentration ( <2 mmol/L) AQS was a accelerator for bacterial azoreduction. However, when the concentration of AQS was more than 5 mmol/L a strong inhibition was occurred. On the other hand, the concentration of AQDS as high as 12 mmol/L the inhibition of azoreduction was still not exhibited, but the effect of acceleration was gradually decreased with the concentration of AQDS increasing (1 mmol/L to 12 mmol/L). 6 azo dyes (Table 1) were tested, all of which had a similar results. These results indicated that the humic substances were not only as a redox mediator during the azoreduction, but also as the competitor for electron from respiration chain. Because the humic substances could be act as the terminal electron acceptor for bacterial anaerobic respiration. AQS and AQDS exhibiting different behaviour on the azoreduction were determined by their standard redox potentials.

[Anaerobic humus respiration by Shewanella cinica D14T].

Experimental results suggested Shewanella cinica D14T is capable of humus respiration utilizing various organic acids and some important environmental pollutants (e.g., toluene. etc) as electron donors and AQS or AQDS as a sole terminal electron acceptor under anaerobic condition. The dissimilatory reduction of 1mmol/L AQDS can couple to the production of enough ATP to support cell growth about 60 generations; The oxidization of electron donors was coupled to the reduction of humus, as reduced humus increased corresponding with increasing of electron donor; The typical inhibitors such as Cu2+ which inhibited Fe-S center, Stigmatellin which was methyl-naphthoquinone model, Dicumarol which inhibited oxidized methyl-naphthoquinone transform to reduced one, Metyrapone which was specific inhibitor for P450 enzyme blocked the humus respiration seriously. These were powerful evidences for humus-respiration by D14.

Autonomous pump against concentration gradient.

Using non-equilibrium molecular dynamics and Monte Carlo methods, we have studied the molecular transport in asymmetric nanochannels. The efficiency of the molecular pump depends on the angle and apertures of the asymmetric channel, the environmental temperature and average concentration of the particles. The pumping effect can be explained as the competition between the molecular force field and the thermal disturbance. Our results provide a green approach for pumping fluid particles against the concentration gradient through asymmetric nanoscale thin films without any external forces. It indicates that pumping vacuum can be a spontaneous process.

Elastic large muscular vessel wall engineered with bone marrow­derived cells under pulsatile stimulation in a bioreactor.

Bone marrow­derived cells (BMCs) have demonstrated their ability to differentiate into multiple cell lineages and may be a promising cell source for vascular tissue engineering. Although much progress has been made in the engineering of small blood vessels (<6 mm in diameter) with biodegradable materials such as polyglycolic acid (PGA), it remains a challenge to engineer large vessels (>6 mm in diameter) due to unsatisfactory biomechanical properties. The present study was to engineered an elastic large vessel wall (6 mm in diameter) using a PGA unwoven fibre scaffold covered with BMCs from canine humeri. The cell­PGA sheet was then loaded into a bioreactor designed for the present study, with dynamic pulsatile culture conditions to mimic the physiological vessel environment. After four weeks of the pulsatile stimuli culture, an elastic vessel wall was formed. Histological analyses demonstrated that layers of smooth muscle­like cells and well­oriented collagenous fibres were evenly oriented in the dynamic group. By contrast, disorganised cells and randomly collagenous fibres were apparent in the static group. Furthermore, the engineered vessel wall in the dynamic group exhibited significantly improved biomechanical properties compared with those in static culture group. The approach developed in the present study was demonstrated to have promising potential to be used for the engineering of large vessel as well as other smooth muscle cell­containing tissues, including bladder, urethral and intestinal tissues.

Studies on the spectroscopic behavior of cryptotanshinone, tanshinone IIA, and tanshinone I.

A comparative study on the spectroscopic behavior of cryptotanshinone (CTan), tanshinone IIA (Tan IIA), and tanshinone I (Tan I) has been investigated, including UV-Vis absorption, low temperature phosphorescence (LTP), low temperature fluorescence (LTF), paper substrate-room temperature phosphorescence (PS-RTP), paper substrate-room temperature fluorescence (PS-RTF) and fluorescence in liquid (LF). The effect of pH on the luminescence intensity is discussed. Lifetime and polarization of the LTP and RTP have been examined with phosphorescence lifetime in the range of 0.6-0.9s and polarization in the range of 0.10-0.27. Analytical characteristics of LF, PS-RTF and PS-RTP of CTan, Tan IIA, and Tan I have been studied.

[Experimental study on constructing small-caliber artery by tissue engineering approach].

OBJECTIVE: To investigate the possibility of constructing small-caliber artery by means of tissue engineering. METHODS: Cell-PGA mixtures were made by separately seeding 1 x 10(7) smooth muscle cells and 5 x 10(6) endothelial cells isolated from neonate umbilicus onto PGA scaffold, the cell-PGA constructs were wrapped around a silicone tube before its implantation subcutaneously to nude mice and the mice were sacrificed in 2 and 6 weeks. The tissue engineered artery (TEA) were examined both grossly and immunohistochemically. RESULTS: The gross appearance of TEA was similar to that of the natural counterparts; histologic and immunohistochemical analyses of the neoformed tissues revealed a typical artery structure, including the presence of EC at the luminal surface and the presence of SMC and collagen in the wall. CONCLUSION: TEA with histology similar to natural vessel can be constructed by tissue engineering.

Differentiation of human hair follicle stem cells into endothelial cells induced by vascular endothelial and basic fibroblast growth factors.

Hair follicle stem cells (HFSCs) possess powerful expansion and multi­differentiation potential, properties that place them at the forefront of the field of tissue engineering and stem cell­based therapy. The aim of the present study was to investigate the differentiation of human HFSCs (hHFSCs) into cells of an endothelial lineage. hHFSCs were expanded to the second passage in vitro and then induced by the addition of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) to the culture medium. The expression levels of endothelial cell (EC)­related markers, including von Willebrand factor (vWF), vascular endothelial cadherin (VE)­cadherin and cluster of differentiation (CD)31, were detected by immunofluorescence staining, flow cytometric analysis and reverse transcription­polymerase chain reaction. The hHFSCs expressed vWF, VE­cadherin and CD31 when exposed to a differentiation medium, similar to the markers expressed by the human umbilical vein ECs. More significantly, differentiated cells were also able to take up low­density lipoprotein. The data of the present study demonstrated that an efficient strategy may be developed for differentiating hHFSCs into ECs by stimulation with VEGF and bFGF. Thus, hHFSCs represent a novel cell source for vascular tissue engineering and studies regarding the treatment of various forms of ischaemic vascular disease.

Human hair follicle stem cell differentiation into contractile smooth muscle cells is induced by transforming growth factor-ß1 and platelet-derived growth factor BB.

Smooth muscle cells (SMCs) are important in vascular homeostasis and disease and thus, are critical elements in vascular tissue engineering. Although adult SMCs have been used as seed cells, such mature differentiated cells suffer from limited proliferation potential and cultural senescence, particularly when originating from older donors. By comparison, human hair follicle stem cells (hHFSCs) are a reliable source of stem cells with multi-differentiation potential. The aim of the present study, was to develop an efficient strategy to derive functional SMCs from hHFSCs. hHFSCs were obtained from scalp tissues of healthy adult patients undergoing cosmetic plastic surgery. The hHFSCs were expanded to passage 2 and induced by the administration of transforming growth factor-ß1 (TGF-ß1) and platelet-derived growth factor BB (PDGF-BB) in combination with culture medium. Expression levels of SMC-related markers, including α-smooth muscle actin (α-SMA), α-calponin and smooth muscle myosin heavy chain (SM-MHC), were detected by immunofluorescence staining, flow cytometry analysis and reverse transcription-polymerase chain reaction (RT-PCR). When exposed to differentiation medium, hHFSCs expressed early, mid and late markers (α-SMA, α-calponin and SM-MHC, respectively) that were similar to the markers expressed by human umbilical artery SMCs. Notably, when entrapped inside a collagen matrix lattice, these SM differentiated cells showed a contractile function. Therefore, the present study developed an efficient strategy for differentiating hHFSCs into contractile SMCs by stimulation with TGF-ß1 and PDGF-BB. The high yield of derivation suggests that this strategy facilitates the acquisition of the large numbers of cells that are required for blood vessel engineering and the study of vascular disease pathophysiology.