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Biomedical subjects

T F Lue

Publications and source records attributed to T F Lue.

At least 73 records · Page 4Linked to original sources
PubMed1998

Effect of simulated birth trauma on the urinary continence mechanism in the rat.

OBJECTIVES: To examine the effect of simulated birth injury in an animal model as part of a study on the pathogenesis of stress urinary incontinence (SUI) and the urinary continence mechanism. METHODS: A balloon was inflated in the vaginas of rats for 4 hours to simulate prolonged labor. The effect on the continence mechanism was assessed by functional, anatomic, biochemical, and histologic examinations. The functional test consisted of placing chili powder or a clipped whisker into the rat's nostrils to induce sneezing. Anatomic measurement of the genital hiatus was performed with a caliper. Serum creatine phosphokinase (CPK) and lactate dehydrogenase (LDH) were measured to examine the value of muscle injury in predicting incontinence. c-Fos immunostaining in the spinal cord was used as a marker of nerve injury. These data were then correlated with histopathologic examination of the urethra and pelvic floor tissues. RESULTS: Four weeks after simulated birth injury, SUI was noted in 19 of 48 experimental rats. The genital hiatus was significantly wider in incontinent rats. The serum CPK and LDH levels were markedly elevated, but no difference was noted between the continent and incontinent rats. All experimental rats showed many c-Fos immunostaining neurons in the L6 to S1 spinal cord segments, but none was seen in control rats. Histologic study revealed a marked decrease of ganglion cells in the neural plexuses posterolateral to the vagina in experimental rats. After 4 weeks, muscle necrosis and degeneration, irregular shape and size of muscle fibers, and a change in the type I/II ratio were prominent features in the levator ani. In the urethra, we noted a significant decline in urethral wall musculature (both smooth and striated) in incontinent rats. CONCLUSIONS: In this novel rat model, simulated birth injury resulted in SUI in a portion of the animals. Pathologic changes in the urethra, pelvic ganglia, and levator muscles seem to be the contributing factors to SUI.

Animals↗
PubMed1998

Possible molecular mechanisms of cryoablation-induced impotence in a rat model.

OBJECTIVES: Cryoablation of the prostate has been reported to induce impotence as a consequence of cavernosal nerve injury. This study is designed to investigate the early and late effects of cavernosal nerve cryoablation on growth factor expression and erectile function in a rat model. METHODS: Forty male rats were divided into two groups (n=20 each). The first group underwent unilateral cavernosal nerve freezing (experimental group). Before their euthanization at 1 and 3 months (10 rats each), erectile function was assessed by electrostimulation of the cavernous nerves. The second group served as the control and was killed at the same time points. Western blot and reverse transcriptase-polymerase chain reaction (RT-PCR) techniques were used to identify protein and gene expression of nerve growth factor (NGF), transforming growth factor-alpha (TGF-alpha), epidermal growth factor (EGF), and insulin-like growth factor-1 (IGF-1) in the rat penis and pelvic ganglia. RESULTS: Electrostimulation of the frozen nerve after 3 months revealed a significantly higher maximal intracavernosal pressure and a shorter latency period than in the 1-month group. At 3 months, immunoblot showed upregulation of NGF, TGF-alpha, and the precursor form of IGF-1 protein expression in the penile tissue; RT-PCR showed downregulation of NGF gene expression in the pelvic ganglia of the frozen side. CONCLUSIONS: The results show that erectile function decreased at 1 month and then partially recovered 3 months after cavernosal nerve freezing. This alteration in erectile function was associated with differential gene and protein expression of the growth factors (NGF, TGF-alpha, EGF, and IGF-1). Further studies are required to elucidate the potential role of these growth factors in the prevention and treatment of cryoablation-induced impotence.

Animals↗
PubMed1998

Histological and ultrastructural alterations in an animal model of Peyronie's disease.

OBJECTIVE: To determine the role of transforming growth factor beta (TGF-beta), one of the cytokines known to induce tissue fibrosis, in the induction of a Peyronie's-like condition, and to produce an animal model for the further study of Peyronie's disease. MATERIALS AND METHODS: Twenty-four adult male Sprague-Dawley rats were divided into two groups: in group 1, different concentrations of cytomodulin, a synthetic heptapeptide with TGF-beta-like activity, were injected into the tunica of each of 18 rats and six rats group 2 received saline injections as a control. The tunical tissues were taken after 3 days, 2 and 6 weeks and were examined histologically using Hart and trichrome stains. Electron microscopy was used to examine the ultrastructural changes in the same tissue samples. RESULTS: There were histological and ultrastructural alterations in 15 of 18 rats in group 1 (cytomodulin-injected), especially in tissue examined after 6 weeks. The most prominent histological changes were chronic inflammatory cellular infiltration, focal and diffuse elastosis, thickening, disorganization and clumping of the collagen bundles. The ultrastructural changes were in the form of densely packed collagen, fragmented and scarce elastic fibres, separation of neuronal fibres by interposing clumps of packed collagen, and perivascular collagen deposition as a part of the reorganization of the interstitial matrix. CONCLUSION: Cytomodulin can induce a Peyronie's-like condition in the rat penis, which may explain the role of TGF-beta in the pathogenesis of Peyronie's disease. With further refinement, such rats may be used as an experimental model for studies of Peyronie's disease.

Animals↗
PubMed1998

Functional rat bladder regeneration through xenotransplantation of the bladder acellular matrix graft.

OBJECTIVE: To determine the decreased antigenicity of the bladder acellular matrix graft (BAMG) through xenotransplantation and to assess the in vivo and in vitro functional properties of the rat urinary bladder thus regenerated. MATERIALS AND METHODS: After partial cystectomy (> 50%), BAMGs prepared from hamster, rabbit and dog urinary bladders were grafted to male and female Sprague-Dawley rats; 10 control rats underwent partial cystectomy only. Urinary storage and voiding function were monitored in 15 animals using a specially designed 'micturition cage' and cystometry. After 4 months, organ-bath studies and histological techniques were used to evaluate bladder regeneration in vitro in the grafted animals. RESULTS: Clinically relevant antigenicity was not evident; no animal died from rejection and all bladder wall components regenerated in all BAMG xenografts. However, the degree and quality of regeneration varied. Muscularization, peak pressure, and bladder capacity were higher in the hamster BAMG-grafted animals, whereas in vitro contractility and compliance were best in the dog BAMG-regenerated bladders. All grafted bladders had significantly better capacity and compliance than the autoregenerated bladders after partial cystectomy alone. CONCLUSIONS: The present in vivo and in vitro studies show that BAMG-augmentation cystoplasty can lead to morphological and functional regeneration of the rat bladder, preserving its low-pressure reservoir function. Because BAMG-regenerated bladders show functional innervation that is similar to normal bladders, they can work in coordination with the host bladder components, thus generating adequate intravesical pressure to produce sustained voiding. The decreased antigenicity makes heterologous BAMG transplants feasible without immunosuppression.

Animals↗
PubMed1998

The correction of congenital penile curvature in young men.

OBJECTIVE: To determine the long-term outcome of multiple parallel plication (MPP) used for correcting congenital penile curvature (with normal urethra and corpus spongiosum), a technique which takes account of the greater force of erection in young men (mean arterial pressure approximately 100 mmHg) than in neonates (approximately 60 mmHg). PATIENTS AND METHODS: All young men who underwent surgical correction of congenital penile curvature using the multiple parallel plication technique between 1992 and 1995 were reviewed, from their charts and by a telephone survey. RESULTS: Ten patients (mean age 22.3 years, SD 4, range 17-29) were identified, all of whom had been followed for more than one year (mean 2.8, SD 1.3, range 1.2-4.8). All 10 patients reported a preoperative curvature of 60-90 degrees; postoperatively, eight of the 10 patients had a successful outcome, based on self-documentation of a straighter penis (eight) and/or satisfactory intercourse (four). Two of these eight requested follow-up plications to correct a residual curvature of 15 degrees. All patients reported feeling the sutures as a small bump on the dorsal aspect of the penis. In one patient this was considered bothersome, but it did not interfere with intercourse. In one patient the curvature recurred 5 weeks after surgery, following intercourse. Another patient was lost to follow-up and could not be evaluated. CONCLUSION: The MPP technique has the following advantages; it entails no dissection of the neurovascular bundle or corpus spongiosum of the tunica, avoiding complications (such as impotence and urethral injury) that may occur with manipulation of these structures; it can be performed under local anaesthesia and can be completed within 30 min; suture cut-through is unlikely because the dorsum (12 o'clock position) of the tunica albuginea is the thickest and hence strongest area of the corporal body; and the straightness of the penis can be determined easily because erection is sustained throughout the procedure. The disadvantages of the MPP technique are; several small bumps from the nonabsorbable sutures may be palpable, and the patient needs to be informed of this preoperatively: aspiration and phenylephrine injection may be required to prevent priapism.

Adolescent↗
PubMed1998

Composition and biomechanical properties of the bladder acellular matrix graft: comparative analysis in rat, pig and human.

OBJECTIVE: To compare the composition and mechanical properties of the newly developed bladder acellular matrix graft (BAMG) with the normal urinary bladder in rat, pig and human. MATERIALS AND METHODS: Rat, pig and human urinary bladders were harvested and divided into control and experimental groups. For the latter, BAMGs were prepared, and light and transmission electron microscopic studies performed. Strips from the normal bladders and the BAMGs (10 in each group) were tested under tension, and the ultimate tensile strength, maximum strain, and elastic modulus were determined from stress/strain curves. RESULTS: Both types I and III collagen, as well as elastic fibres, were observed as major components of the matrix scaffold. There were more collagen type I fibres in the rat than in the pig and human BAMGs, whereas the pig, and particularly the human, both showed higher levels of type III collagen and elastic fibres. These different matrix scaffold patterns were confirmed by electron microscopy. Results from biomechanical testing showed no significant differences for strength, strain or elastic modulus between BAMG and control bladder strips, except in the rat where the maximum strain values were significantly lower. CONCLUSION: There are variations in the acellular matrix structure with similar biomechanical properties between the BAMG and the normal urinary bladder in three different species. These results may underscore the potential of the BAMG. Furthermore, this in vitro model provides a suitable method to study the mechanical properties of the urinary bladder and may serve as a diagnostic tool for various investigations.

Animals↗
PubMed1998

Analysis and mapping of plastin phosphorylation.

Plastins (fimbrins) are a family of three tissue-specific actin-binding proteins. Both L- and T-plastin have been shown to be involved in cytoskeletal reorganization in signal-transduction pathways. Phosphorylation of the leukocyte-specific L-plastin plays important roles in regulating L-plastin function and leukocyte activation. We created L-plastin mutants, changing each of the Ser-5 and Ser-7 residues to alanine. Expression of either mutant in WI38VA13 fibroblasts did not result in phosphorylation. The transfected wildtype L-plastin was phosphorylated in WI38VA13, but its two-dimensional gel pattern suggested that it was phosphorylated on one residue, whereas the endogenous L-plastin in leukocytes was likely phosphorylated on two residues. The nonleukocyte-specific T-plastin, which has an equivalent Ser-7 residue, was not phosphorylated in T-plastin-expressing fibroblasts or in transfected leukocytes. Expression of CK-IIalpha, the catalytic subunit of casein kinase II, resulted in changes of the protein expression profile in leukocytes but not the phosphorylation status of L- or T-plastin. Phorbol myristate acetate induced L-plastin phosphorylation in both leukocytes and fibroblasts, lending support to the view that protein kinase C is the likely candidate kinase for L-plastin phosphorylation.

Amino Acid Sequence↗
PubMed1998

Health outcomes variables important to patients in the treatment of erectile dysfunction.

PURPOSE: Erectile dysfunction is underreported and the rate of noncompliance with therapy is high. The National Institutes of Health Consensus Conference on Impotence endorsed the need for outcomes research of the various approaches to treatment. The purpose of our exploratory study was to begin that process through identification of erectile dysfunction treatment outcomes variables that are important to men. MATERIALS AND METHODS: The study format consisted of focus group meetings. The goal of the discussions was to identify variables that are important to men when choosing among and judging the success of treatment modalities for erectile dysfunction. After discussion participants were individually asked to identify the 10 variables that they considered important and rank the 5 most important variables. An importance score reflecting group consensus was calculated for each variable. RESULTS: Success and negative outcomes were identified as the 2 most important outcomes variables for all treatment modalities. Participants defined success in several ways. The most important measures of success were cure, pleasure and partner satisfaction. Negative outcome was defined as negative consequences associated with treatment. CONCLUSIONS: The focus group discussion assisted in identifying and defining outcomes variables important to patients with erectile dysfunction. We believe that understanding issues and outcomes important to such patients is necessary to increase compliance with treatment and, thereby, increase the success of treatment of this widespread condition.

Adult↗
PubMed1998

The effect of surgical trauma on rat tunica albuginea.

PURPOSE: Increased TGF-beta1 protein expression in tunica albuginea has been found to be associated with Peyronie's disease. The present study is designed to investigate whether surgical trauma induces TGF-beta up-regulation and histological changes in rat penis. MATERIALS AND METHODS: Thirty-two adult male Sprague-Dawley rats were divided into two groups. The first group (n=24) underwent incision and suture repair of the tunica albuginea of the penis. The second group (n=8) received sham surgery (incision of the penile skin and underlying fascia) as the control group. The trauma-induced group was divided into four subgroups in which the rats were euthanized at 6 hours (n=6), 1 day (n=6), 3 days (n=6), and 8 weeks (n=6). Two sham-operated (control) animals were also euthanized at each of the above time points. All tunical tissues from the trauma-induced and sham-operated rats were collected and examined histologically using Trichrome and Hart elastic fiber stain. Electron microscopy was used to study the ultrastructural changes of both trauma induced and control specimens. Western blotting technique was performed to study TGF-beta protein expression in both experimental and sham-operated groups. RESULTS: Tissue edema and hemorrhage between collagen bundles are noted in the experimental groups after 6 hours, 1 day and 3 days. At 8 weeks the most prominent changes observed were inflammatory cellular infiltration and disorganization of the collagen bundles. In the control group the tunica albuginea retains normal wavy regular appearance in all rats. This histological analysis is similar to the reported description of histological features of the acute phase of Peyronie's disease. Electron microscopy showed packed collagen bundles in the trauma-induced group with normal appearing elastic fibers. No abnormal change was detected in the control group. Immunoblot results revealed remarkable TGF-beta1 protein expression in 1, 5, 3, and 0 rats of trauma induced subgroups after 6 hours, 1 day, 3 days, and 8 weeks respectively. No TGF-beta1 protein expression in any rats in the control group was detected. No significant TGF-beta2 or TGF-beta3 protein expression was observed either in the trauma induced group or in the control group. CONCLUSION: Trauma can induce histological changes similar to the acute phase of Peyronie's disease but not the overt picture of the chronic phase of Peyronie's disease. It can also result in an early but transient up-regulation of TGF-beta1 protein expression in the rat penis. We conclude that surgical incisional trauma does not result in Peyronie's disease-like changes in the tunica.

Animals↗
PubMed1998

In vitro functional properties of the rat bladder regenerated by the bladder acellular matrix graft.

PURPOSE: To assess the response of rat urinary bladder regenerated by the homologous bladder acellular matrix graft (BAMG) to in vitro electrical and pharmacologic stimuli. MATERIALS AND METHODS: In Sprague-Dawley rats, partial cystectomy (>50%) was performed, followed by BAMG augmentation cystoplasty. After 4 months, organ bath studies of tissue strips in 10 were used to compare the contractility of the BAMG regenerates and the corresponding host detrusor smooth muscle. RESULTS: The BAMG regenerates exhibited contractile activity to electrical field stimulation and a qualitatively identical pattern of response to muscarinic, purinergic, alpha- and beta-adrenergic drug administration and nitric oxide. At 4 months after surgery, the maximum forces of contraction of the BAMG regenerates to carbachol stimulation amounted to close to 80% of the host bladder response. With electrical field stimulation, they equaled 44% and 62% of the host bladder response after 2.5 and 4 months, respectively. Histological and immunohistochemical studies confirmed the presence of receptors for neurotransmitters that these functional in vitro studies implied. CONCLUSIONS: The present study provides further evidence that augmentation cystoplasty with the BAMG leads to functional regeneration of the rat bladder detrusor smooth muscle.

Animals↗
PubMed1998

Venous patch graft for Peyronie's disease. Part I: technique.

PURPOSE: We describe our technique of plaque incision and venous patch grafting to correct complex penile deformity associated with Peyronie's disease. MATERIALS AND METHODS: Graft material is obtained from the lower and upper saphenous and deep dorsal veins. The configuration, size and number of tunical incisions depend on the site and size of the lesion. In most cases an H-shaped tunical incision is adequate to release the contracture. With the aid of a vascular stapler several vein segments can be assembled easily into 1 piece to cover the defect. RESULTS: The incidence of penile shortening and erectile dysfunction, following other corrective procedures, is lessened with the tunical incision and venous grafting technique. CONCLUSION: The venous graft provides an anatomical and functional tunical substitute. Results in correction of Peyronie's disease are highly encouraging.

Humans↗
PubMed1998

Venous patch graft for Peyronie's disease. Part II: outcome analysis.

PURPOSE: We evaluate the results of tunical incision and venous patch grafting for correcting penile deformity in Peyronie's disease. MATERIALS AND METHODS: In 113 [corrected] patients with symptoms of Peyronie's disease for more than a year indications for surgery included penile shortening, persistent pain, severe curvature, penile narrowing or indentation and/or failure of previous surgery. Preoperative evaluation included determination of patient and partner expectation, potency status, circumcision status, measurement of penile length (short and long side) and saphenous vein, and color duplex ultrasonography to evaluate possible accessory vascular communication. Patients underwent plaque incision and venous patch grafting. The configuration, size and number of tunical incisions depended on the size and shape of the lesion. Lower and upper saphenous, and deep dorsal veins served as the graft materials. Postoperative followup was as long as 18 months. RESULTS: In 96% of patients the penis became straight, while residual curvature was 30 degrees in 3% and 15 degrees in 1%. In 94% of patients narrowing and indentation were absent and in 83% penile length was the same or longer postoperatively. Of the patients who were potent preoperatively 88% experienced the same or better erectile quality after surgery. In 10% of cases a change in sensation occurred lasting longer than 6 months. Overall satisfaction was expressed by 92% of men who believed that surgery improved the psychological state as well as the relationship with the partner. CONCLUSIONS: The results are satisfactory and this procedure offers a reasonable solution for correction of Peyronie's disease.

Adult↗
PubMed1998

Effect of cavernous nerve freezing on protein and gene expression of nitric oxide synthase in the rat penis and pelvic ganglia.

PURPOSE: Cryoablation of the prostate has been reported to induce impotence as a result of cavernous nerve injury. This study is designed to investigate the early and late effects of cavernosal nerve cryoablation on erectile function and nitric oxide synthase (NOS) containing nerves in the rat penis and pelvic ganglia. MATERIALS AND METHODS: Forty male rats were divided into two groups (n = 20 each). The first group underwent unilateral cavernous nerve freezing (experimental group). Before their euthanization at 1 and 3 months (10 rats each), erectile function was assessed by electrostimulation of the cavernous nerves. The second group served as a control and was euthanized at the same time points. Nicotinamide adenine dinucleotide phosphate (NADPH) diaphorase staining was used to identify NOS in the penile nerve fibers of the mid-shaft segment and neurons of the pelvic ganglia. Western blot, RT-PCR and immunostaining techniques were used to identify protein and gene expression of eNOS, iNOS, and nNOS. RESULTS: One month after unilateral cavernosal nerve freezing, there was a significant decrease in NOS-containing nerve fibers in the dorsal and intracavernosal nerves and ipsilateral pelvic ganglia as compared with the intact side. At three months, the number of NOS containing nerve fibers in the above-mentioned nerves and ganglia showed a significant increase on the frozen side but not the intact side. Electrostimulation of the frozen nerve after three months revealed a significantly higher maximal intracavernosal pressure and a shorter latency period than the one-month group. At three months, immunoblot showed up-regulation of eNOS and nNOS protein expression in the penile tissue. RT-PCR showed up-regulation of nNOS gene expression in the pelvic ganglia of the frozen side. Immunostaining confirmed the results of western blot and showed significant increase of the nNOS positive staining in the frozen side of the penile tissue after three months. There was no difference in iNOS after three months between both sides. Our repeated eNOS immunostaining was not successful. CONCLUSIONS: The results show that intracavernous pressure response to neurostimulation markedly decreased at one month and then partially recovered three months after cavernosal nerve freezing. A similar pattern of changes of the NOS-containing nerve fibers in dorsal nerves, intracavernosal nerves, and pelvic ganglia were observed. This alteration in erectile function and NOS-containing nerves was associated with differential gene and protein expression of the three subtypes of NOS.

Animals↗
PubMed1998

Expression of TGF-beta-1 mRNA and ultrastructural alterations in pharmacologically induced prolonged penile erection in a canine model.

PURPOSE: Transforming growth factor beta (TGF-beta) is known to induce fibrosis. Our objective was to study the role of TGF-beta as a possible mediator of fibrosis that may follow prolonged penile erection. MATERIALS AND METHODS: Prolonged penile erection was induced in seven adult male mongrel dogs by intracavernosal injection of papaverine into one of the corpora cavernosa while the other was used as a control. Intracavernosal pressure measurements were carried out prior to administration of papaverine and at the end of the procedure. Penile tissue was collected from anesthetized animals prior to euthanasia for histological and electron microscopic (EM) studies. RT-PCR was carried out for detection of mRNA on same tissue samples. RESULTS: The light microscopy showed stasis of blood in the cavernosal sinusoids. EM studies revealed sporadic endothelial defects, loss of plasma membrane integrity and cytoplasmic condensation. There was expression of TGF-beta1 mRNA in 66.7% of the experimental group compared with 16.7% of the control group. CONCLUSIONS: Pharmacologically induced low flow prolonged penile erection in canine models is associated with histomorphological changes in relatively short periods of time, suggesting that early therapeutic intervention is desirable. The gene expression for TGF-beta1 may be a mediator of fibrosis; therefore the use of anti-TGF-beta agents presents a possible tool for therapeutic intervention.

Animals↗
PubMed1998

Electroacupuncture decreases c-fos expression in the spinal cord induced by noxious stimulation of the rat bladder.

PURPOSE: The present study investigated the effects of noxious stimulation of the lower urinary tract on neuronal fos protein expression in the spinal micturition center of rats and also examined the effects of electroacupuncture (EA) on fos expression induced by noxious stimulation of the lower urinary tract. MATERIALS AND METHODS: Experiments were conducted on 21 female Sprague-Dawley rats divided into four groups. Group 1 rats (n = 5) served as normal controls. Group 2 rats (n = 5) received EA at the Sanyinjiao acupoint. Group 3 animals (n = 6) were catheterized through the urethra and instilled with 1% acetic acid, and group 4 (n = 5) animals received EA 1 hour before 1% acetic acid instillation. All animals underwent arterial perfusion, laminectomy, and spinal cord removal. Spinal cords were sectioned and processed for immunohistochemical staining for fos protein. RESULTS: No fos protein was detected in any spinal neurons in normal control animals, and either none or few (0 to 4 cells/section) fos-immunoreactive (fos-IR) cells were seen in animals treated with EA. Noxious stimulation of the lower urinary tract with 1% acetic acid drastically increased the number of fos-IR neurons (30 to 127 cells/L6 section, mean 76.17+/-13.98; 28 to 77, cells/S1 section, mean 59+/-8.30; 7 to 35 cells/S2 section, mean 19.83+/-4.10). However, EA administered 1 hour before 1% acetic acid instillation significantly decreased the number of fos-IR neurons resulting from chemical irritation (0 to 50 cells/L6 section, mean 19.8+/-9.33; 0 to 47 cells/S1 section, mean 13.2+/-9.12; 0 to 37 cells/S2 section, mean 13.6+/-7.31). CONCLUSIONS: Our study demonstrates that bladder instillation with 1% acetic acid induces fos protein expression in the spinal micturition center of the rat and that electroacupuncture can reduce this expression. These results suggest a link between electroacupuncture and reduction in spinal neuronal cell activity.

Acetic Acid↗
PubMed1998

Peyronie's disease.

The pathogenesis of Peyronie's disease is still not well understood. As a result, the treatment of Peyronie's disease remains a dilemma and new therapies continue to evolve. This article discusses present understanding, controversy and new discoveries related to this condition.

Journal Article↗
PubMed1998

Venous grafting for the correction of penile curvature in Peyronie's disease.

A number of procedures have been developed to correct penile deformity secondary to Peyronie's disease. In many cases, tunica-shortening procedures have had reasonable success. The most popular of these are tunical plication and Nesbit's wedge resection. However, these procedures shorten the penis and do not correct the hourglass deformity. Tunica-lengthening by using autologous or synthetic materials has been reported with varying success. However, notable shortcomings including graft contracture, recurrence, and impotence have been reported. This review describes our experience with tunica incision and venous grafting.

Journal Article↗