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S Kröger

Publications and source records attributed to S Kröger.

At least 19 recordsLinked to original sources

Molecular diversity of the dystrophin-like protein complex in the developing and adult avian retina.

Mutations in dystrophin cause muscular dystrophy but also affect the CNS, including information processing in the retina. To better understand the molecular basis of these CNS deficits, we analyzed the molecular composition and developmental appearance of dystrophin and of the dystrophin-associated protein complex (DPC) in the embryonic and adult avian retina. We detected a concentration of the DPC at the vitreal border and in the outer plexiform layer of the adult retina. At both locations the complex had a different molecular composition and different developmental expression pattern. At the vitreal border, the complex was composed of utrophin, alpha-dystrobrevin-1, and dystroglycan, and was present at all stages of retinal development even before neurogenesis and gliogenesis. On the other hand, the complex in the outer plexiform layer consisted of dystrophin, beta-dystrobrevin and dystroglycan. The distribution of this complex changed from a diffusely distributed to an aggregated form during development concomitant with synapse formation in the outer plexiform layer. Solubilization of the retinal extracellular matrix by intravitreal injection of collagenase resulted in a redistribution of the complex at the retinal vitreal border but had no influence on the distribution of the dystrophin-associated proteins in the outer plexiform layer. These results demonstrate two types of dystrophin-like complexes in the chick retina with differential molecular compositions, different anchorage to the extracellular matrix, and different developmental expression patterns, suggesting distinct functions for the DPC at both locations.

Aging↗

An alternative amino-terminus expressed in the central nervous system converts agrin to a type II transmembrane protein.

Agrin is a basal lamina-associated heparansulfate proteoglycan that is a key molecule in the formation of the vertebrate neuromuscular junction. The carboxy-terminal part of agrin is involved in its synaptogenic activity. The amino-terminal end of chick agrin consists of a signal sequence, required for the targeting of the protein to the secretory pathway, and the amino-terminal agrin (NtA) domain that binds to basal lamina-associated laminins. The cDNA encoding rat agrin lacks this NtA domain and instead codes for a shorter amino-terminal end. While the NtA domain is conserved in several species, including human, sequences homologous to the amino-terminus of rat agrin have not been described. In this work, we have characterized these amino-terminal sequences in mouse and chick. We show that they all serve as a noncleaved signal anchor that immobilizes the protein in a N(cyto)/C(exo) orientation in the plasma membrane. Like the secreted form, this transmembrane form of agrin is highly glycosylated indicative of a heparansulfate proteoglycan. The structure of the 5' end of the mouse agrin gene suggests that a distinct promoter drives expression of the transmembrane form. Agrin transcripts encoding this form are enriched in the embryonic brain, particularly in neurons. To our knowledge, this is the first example of a molecule that is synthesized both as a basal lamina and a plasma membrane protein.

Agrin↗

Decreased glomerular expression of agrin in diabetic nephropathy and podocytes, cultured in high glucose medium.

AIM: A decrease in glomerular heparan sulfate (HS) proteoglycan (PG), without apparent decrease in HSPG core protein expression, has been reported to occur in diabetic nephropathy (DN). In most studies however, agrin, the major HSPG core protein in the glomerular basement membrane, has not been studied. This prompted us to study the glomerular expression of agrin in parallel to the expression of HS-glycosaminoglycans (GAG) in biopsies of patients with DN. Furthermore, the influence of glucose on agrin production in cultured podocytes and the expression of agrin in fetal kidneys was investigated. METHODS: Cryostat sections of renal biopsies from patients with DN (n = 8) and healthy controls (HC, n = 8), were stained for agrin and HS-GAG. Sections of fetal kidneys were double stained for agrin and CD35 or CD31. Stainings were performed by indirect immunofluorescence (IIF). The production of agrin by cultured human podocytes was tested by ELISA and IIF. RESULTS: The expression of agrin, detected by AS46, was significantly reduced in biopsies from patients with DN compared to HC (p < 0.01). Similar findings were observed when monoclonal antibody JM72 was used (p < 0.05). In addition, a significant reduction in the glomerular expression of HS-GAG was detected with JM403 in these patients (p < 0.01). Agrin is expressed in cultured podocytes, the expression hereof was reduced when the cells were cultured in the presence of 25 mM D-glucose (p < 0.01). In biopsies of human fetal kidneys, glomerular expression of agrin coincided with the expression of CD31. In early stages of glomerular differentiation there was a strong staining for agrin and CD31 while CD35 was only slightly positive. CONCLUSIONS: Our data argue against a selective dysregulation in HSPG sulfation in DN, but suggest a pivotal role for hyperglycemia in the downregulation of agrin core protein production.

Aged↗

[Introduction of a quality management system compliant with DIN EN 9001:2000 in a university department of nuclear medicine].

In 1995, the management of the University Clinic Hamburg-Eppendorf proposed to establish a total quality assurance (QA) system. A revised QA-system has been introduced stepwise in the department of nuclear medicine since 1997, and certification was achieved in accordance with DIN EN ISO 9001:2000 on February 14, 2001. The QA-handbook is divided into two parts. The first part contains operational (diagnostic and therapeutic) procedures in so-called standard operating procedures (SOP). They describe the indication of procedures as well as the competences and time necessary in a standardized manner. Up to now, more than 70 SOPs have been written as a collaborative approach between technicians and physicians during daily clinical routine after analysing and discussing the procedures. Thus, the results were more clearly defined processes and more satisfied employees. The second part consists of general rules and directions concerning the security of work and equipment as well as radiation protection tasks, hygiene etc. as it is required by the law. This part was written predominantly by the management of the department of nuclear-medicine and the QA-coordinator. Detailed information for the patients, documentation of the work-flows as well as the medical report was adopted to the QM-system. Although in the introduction phase of a QA-system a vast amount of time is necessary, some months later a surplus for the clinical workday will become available. The well defined relations of competences and procedures will result in a gain of time, a reduction of costs and a help to ensure the legal demands. Last but not least, the QA-system simply helps to build up confidence and acceptance both by the patients and the referring physicians.

Documentation↗

Dissociation of the dystroglycan complex in caveolin-3-deficient limb girdle muscular dystrophy.

Limb girdle muscular dystrophy is a group of clinically and genetically heterogeneous disorders inherited in an autosomal recessive or dominant mode. Caveolin-3, the muscle-specific member of the caveolin gene family, is implicated in the pathogenesis of autosomal dominant limb girdle muscular dystrophy 1C. Here we report on a 4-year-old girl presenting with myalgia and muscle cramps due to a caveolin-3 deficiency in her dystrophic skeletal muscle as a result of a heterozygous 136G-->A substitution in the caveolin-3 gene. The novel sporadic missense mutation in the caveolin signature sequence of the caveolin-3 gene changes an alanine to a threonine (A46T) and prevents the localization of caveolin-3 to the plasma membrane in a dominant negative fashion. Caveolin-3 has been suggested to interact with the dystrophin-glycoprotein complex, which in striated muscle fibers links the cytoskeleton to the extracellular matrix and with neuronal nitric oxide synthase. Similar to dystrophin-deficient Duchenne muscular dystrophy, a secondary decrease in neuronal nitric oxide synthase and alpha-dystroglycan expression was detected in the caveolin-3-deficient patient. These results implicate an important function of the caveolin signature sequence and common mechanisms in the pathogenesis of dystrophin-glycoprotein complex-associated muscular dystrophies with caveolin-3-deficient limb girdle muscular dystrophy.

Amino Acid Substitution↗

Distribution of the integrin beta 1 subunit on radial cells in the embryonic and adult avian retina.

The distribution of the beta1 integrin subunit was investigated in the developing and adult chick retina at the light and electron microscopic levels, using two different monoclonal antibodies. Western blotting revealed a single band with a molecular weight of approximately 130 kDa in the retina and in a number of other tissues, indicating the specificity of the antibodies. In the retina, immunoreactivity was detected on radial cells spanning the entire width between the pigment epithelium and the vitreal border. These cells were undifferentiated neuroepithelial cells at early stages and radial Müller glial cells at later stages of development. At all stages, the beta1 subunit was concentrated at the vitreal border of the retina around the inner limiting membrane. Mechanical isolation of the inner limiting membrane, as well as immunoelectron microscopy, demonstrated that this immunoreactivity was due to a concentration of the beta1 subunit in the endfeet of neuroepithelial and Müller glial cells. Injection of collagenase into the vitreous of live embryos, a procedure that selectively removes the inner limiting membrane, but does not proteolytically degrade the integrin protein, resulted in a redistribution of the integrin immunoreactivity, demonstrating that the integrity of the basal lamina is required for the maintenance of the concentration of the beta1 subunit in the endfeet. These results suggest a role for the beta1 subunit-containing integrin heterodimers in the adhesion of neuroepithelial and Müller glial cells to extracellular matrix components of the inner limiting membrane, possibly stabilizing the radial morphology of these cells.

Aging↗

The neurobiology of duchenne muscular dystrophy: learning lessons from muscle?

Several forms of inherited muscular dystrophy are associated with brain abnormalities and cognitive impairment. One of the most common and severe of these diseases is Duchenne muscular dystrophy (DMD). Dystrophin, the product of the DMD gene, is found in neurones, where it is associated with the postsynaptic membrane. Cognitive impairment in individuals with DMD is thought to be due to an abnormality in the neuronal membrane that is caused by lack of dystrophin. Recent experimental evidence has provided valuable clues in our understanding of the complex molecular neurobiology of muscular dystrophy.

Animals↗

Expression of EMAP II in the developing and adult mouse.

Endothelial monocyte-activating polypeptide II (EMAP II) is a chemoattractant for monocytes and granulocytes. EMAP II is translated as a precursor protein, proEMAP II, and is proteolytically cleaved to become the mature, biologically active cytokine. In this study we show that the EMAP II mRNA and the EMAP II precursor protein are constitutively expressed by all cell types analyzed in vitro, whereas the mature cytokine is only present in the supernatant of apoptotic cells. During mouse embryogenesis we found widespread expression of the EMAP II mRNA with transcripts being abundant in areas of tissue remodeling, where a large number of apoptotic cells could be detected by TUNEL staining. In the adult mouse, strong expression of the EMAP II mRNA is restricted to the brain, testis and thymus. Interestingly, prominent signals for EMAP II mRNA are found in local correlation with sites of apoptosis in thymus and testis but not in the brain. We propose that during development, the generation and release of the mature EMAP II may provide a mechanism for the recruitment of phagocytic cells to sites of programmed cell death. In the adult brain, the generation of mature EMAP II may contribute to the recruitment of monocytes and the immunosurveillance of this tissue.

Animals↗

FDG PET detection of unknown primary tumors.

UNLABELLED: The management of patients presenting with metastases of unknown primary origin remains a clinical challenge despite a large variety of imaging modalities. The aim of this study was to evaluate FDG PET in detecting the sites of primary cancer in these patients. METHODS: Fifty-three patients with metastatic cervical adenopathy (n = 44) or extracervical metastases (n = 9) of unknown primary origin were included after extensive but inconclusive conventional diagnostic work-up. Patients received 370 MBq FDG (10 mCi) intravenously, and whole-body images were acquired at 60 min after injection. Clinical, surgical, and histopathologic findings and complete correlative imaging were used to assess the results. RESULTS: In 27 of 53 patients FDG PET showed focal tracer accumulations corresponding to potential primary tumor sites located in the lungs (n = 12), the palatine tonsil (n = 5), the salivary glands (n = 2), the nasopharynx (n = 1), the oropharynx (n = 3), the maxillary sinus (n = 1), and the larynx (n = 1). Moreover, in 2 patients FDG PET revealed lesions suspected to be tumors in the breast and the ileocolonic area. In 20 (37.8%) of these 53 patients FDG PET was true-positive, identifying the primary tumor in the lungs (n = 10), the head and neck region (n = 8), the breast (n = 1), and the ileocolonic area (n = 1). In 6 of 27 patients FDG PET was false-positive, predominantly identifying suspicious areas in the palatine tonsil (n = 3). One patient denied further diagnostic work-up after PET; thus, positive PET could not be evaluated. In 26 of 53 patients PET did not reveal lesions suspected to be the primary. However, primary tumors were not found in these patients at clinical follow-up. CONCLUSION: FDG PET is a valuable diagnostic tool in patients with cancer of unknown primary because it imaged unknown primary tumors in about one third of all patients investigated. In addition, FDG PET assists in both guiding biopsies for histologic evaluation and selecting the appropriate treatment protocols for these patients.

Adult↗

Synapse formation and agrin expression in stratospheroid cultures from embryonic chick retina.

Stratospheroids are three-dimensional cellular spheres which develop in vitro through the proliferation and differentiation of retinal neuroepithelial precursor cells. We investigated synapse formation in stratospheroids by analyzing the development of aggregates of synapse-associated molecules and of electron microscopically identifiable synaptic specializations. Our results show that the first aggregates of the GABA(A) receptor, the glycine receptor, and gephyrin appear in the inner plexiform layer after 8 days in culture simultaneously with the development of the first active zones and postsynaptic densities. In contrast, presynaptic molecules including synaptophysin could be detected in the inner plexiform layer before synaptogenesis, suggesting functions for these molecules in addition to neurotransmitter exocytosis at mature synapses. Similar to the retina in vivo, synapses were not found in the nuclear layers of stratospheroids. We also analyzed the isoform pattern, expression, and distribution of the extracellular matrix molecule agrin, a key regulator during formation, maintenance, and regeneration of the neuromuscular junction. In stratospheroids, several agrin isoforms were expressed as highly glycosylated proteins with an apparent molecular weight of approximately 400 kDa, similar to the molecular weight of agrin in the retina in vivo. The expression specifically of the neuronal isoforms of agrin was concurrent with the onset of synaptogenesis. Moreover, the neuronal agrin isoforms were exclusively found in the synapse-containing inner plexiform layer, whereas other agrin isoforms were associated also with the inner limiting membrane and with Müller glial cells. These results show that synapse formation is very similar in stratospheroids and in the retina in vivo, and they suggest an important role for agrin during CNS development.

Agrin↗

Imprinted polymer-based sensor system for herbicides using differential-pulse voltammetry on screen-printed electrodes.

A sensor system for the herbicide 2,4-dichlorophenoxy-acetic acid has been developed based on specific recognition of the analyte by a molecularly imprinted polymer and electrochemical detection using disposable screen-printed electrodes. The method involves a competitive binding step with a nonrelated electrochemically active probe. For batch binding assays, imprinted polymer particles are incubated in suspension with the analyte and the probe, followed by centrifugation and quantification of the unbound probe in the supernatant. Two different compounds, namely 2,4-dichlorophenol and homogentisic acid, were tested as potential electroactive probes. Both compounds could be conveniently detected by differential-pulse voltammetry on screen-printed, solvent-resistant three-electrode systems having carbon working electrodes. Whereas 2,4-dichlorophenol showed very high nonspecific binding to the polymer, homogentisic acid bound specifically to the imprinted sites and thus allowed calibration curves for the analyte in the micromolar range to be recorded. An integrated sensor was developed by coating the imprinted polymer particles directly onto the working electrode. Following incubation of the modified electrode in a solution containing the analyte and the probe, the bound fraction of the probe is quantified. This system provides a cheap, disposable sensor for rapid determination of environmentally relevant and other analytes.

Electrochemistry↗

Dystrophin and beta-dystroglycan in photoreceptor terminals from normal and mdx3Cv mouse retinae.

Mutations in the dystrophin gene cause muscular dystrophy as well as cognitive impairments, including an abnormal dark-adapted electroretinogram. To investigate the basis for the ocular phenotype, we analysed dystrophin and the dystrophin-associated protein beta-dystroglycan in retinae from mdx3Cv mice. This strain has a mutation in the dystrophin gene and abnormalities in the electroretinogram which are similar to those of muscular dystrophy patients. Despite an overall reduction of all dystrophin isoforms and of beta-dystroglycan in retinal tissue from mutant mice, we observed no apparent change in the histotypic layering of the retina, or in the ultrastructure of several specific cell types, including rods and cones. In retinae from wild type and mdx3Cv mice, dystrophin and beta-dystroglycan were concentrated in small extensions of rod and cone photoreceptor terminals protruding into the outer plexiform layer. Beta-dystroglycan but not dystrophin was also clustered around the inner limiting membrane and the capillary basal laminae. While the labelling pattern around the basal laminae was not altered in the mutant mice, we found that the area as well as the intensity of the dystrophin and beta-dystroglycan immunoreactivity associated with the terminals of rod photoreceptors were severely reduced. The same parameters were much less affected in cone terminals. These results show, that dystrophin and beta-dystroglycan are differentially distributed in the retina, and that a severe reduction of dystrophin has no gross effect on retinal structure, but could influence intraretinal signalling at the level of the photoreceptor terminals. Moreover, the mutation in mdx3Cv mice has a selective effect on rods, providing an explanation for the altered electroretinogram.

Animals↗

Expression, distribution and ultrastructural localization of the synapse-organizing molecule agrin in the mature avian retina.

At the vertebrate neuromuscular junction the extracellular matrix molecule agrin is responsible for the formation, maintenance and regeneration of most if not all postsynaptic specializations. Several agrin isoforms are generated by alternative splicing which differ in their function and which are all expressed in the CNS. To analyse the role of agrin in the CNS, we investigated the expression and ultrastructural localization of agrin in the posthatched chick retina. In situ hybridization revealed the presence of agrin mRNA in all cellular layers of the mature retina, indicating that most if not all major retinal cell types synthesize agrin. Pan-specific as well as isoform-specific antiagrin antisera stained the optic fibre layer and the outer plexiform layer. However, only the pan-specific antiserum additionally stained the inner limiting membrane. Immunoelectron microscopy showed that in the optic fibre layer agrin was associated with ganglion cell axons and that at least part of this agrin corresponds to a neuronal isoform of agrin. In the outer plexiform layer, agrin was localized in the cleft between the photoreceptor terminals and the invaginating horizontal and bipolar cell dendrites. In the synapse-containing inner plexiform layer both antisera revealed punctate immunoreactivity. This staining corresponded to agrin concentrated in the synaptic cleft of conventional synapses as determined by preembedding immunoelectron microscopy. Agrin is thus concentrated at mature interneuronal synapses as it is at the neuromuscular junction, consistent with a role of agrin during formation and/or maintenance of synapses in the CNS.

Agrin↗

[F-18 FDG PET for detection of occult primary tumor in patients with lymphatic metastases of the neck region].

BACKGROUND: The occult primary tumor remains an important diagnostic problem in patients with lymph node metastases despite a large variety of imaging modalities. Therefore, the aim of our study was to evaluate the impact of F-18-FDG-positron-emission-tomography (PET) in these patients. METHODS: The study group included a total of 28 patients aged 39 to 84 years with cervical lymph node metastases of a histologically confirmed squamous cell carcinoma (n = 24) or an undifferentiated carcinoma (n = 4). All patients received 370 MBq F-18-FDG intravenously, and whole-body images were acquired at 60 min p.i. with an ECAT EXACT 47 (921) (Siemens, CTI). All lesions were evaluated either by histology or by CT/MRI. RESULTS: In 12 patients PET did not reveal suspected tumorous lesions. In contrast in 16/28 patients PET showed focal tracer accumulations corresponding to potential primary tumor sites located in the lungs (n = 7), in the region of the tonsilla palatina (n = 5), in the submandibular gland (n = 1), in the nasopharynx (n = 1), in the larynx (n = 1), or at the base of the tongue (n = 1). In nine out of these 16 patients, the primary was confirmed in the lungs in 5 patients, in the larynx, at the base of the tongue, in the nasopharynx, and the tonsilla palatina in one patient each, respectively. Moreover, in 6 out of 16 patients, PET was false-positive. One patient refused further evaluation of PET findings. CONCLUSIONS: In approximately one third of all patients, PET detected the primary tumor site, which significantly influenced further therapeutic procedures. Thus, F-18-FDG-PET may be a valuable diagnostic tool in the detection of the primary tumor in patients with cervical lymph node metastases.

Adult↗

Salivary gland protection by S-2-(3-aminopropylamino)-ethylphosphorothioic acid (amifostine) in high-dose radioiodine treatment: results obtained in a rabbit animal model and in a double-blind multi-arm trial.

Since differentiated thyroid cancer has an excellent prognosis, reduction of long-term side effects of high-dose radioiodine treatment (HD-RIT), i.e. salivary gland impairment is important. Thus, radioprotective effects of amifostine were studied. Salivary gland function was quantified by scintigraphy both in rabbits and patients. Fifteen rabbits were studied prior to and up to 6 months after HD-RIT applying 2 GBq 131I. Ten animals received 200 mg/kg amifostine prior to HD-RIT, and five served as controls. Animals were examined histopathologically. Fifty patients with differentiated thyroid cancer were evaluated prospectively prior to and 3 months after HD-RIT with either 3 or 6 GBq 131I in a double-blind, placebo-controlled study. Twenty-five patients were treated with 500 mg/m2 amifostine intravenously prior to HD-RIT, and 25 patients receiving physiological saline solution served as controls. Complete ablation of the thyroid was achieved in all rabbits four weeks after HD-RIT. In control rabbits 6 months after HD-RIT parenchymal function was reduced significantly (p < 0.0001) by 75.3 +/- 5.3% and 53.6 +/- 17.4% in parotid and submandibular glands, respectively. In contrast, in amifostine-treated rabbits parenchymal function was not significantly reduced. Histopathologically, marked lipomatosis was observed in control animals but was negligible in amifostine-treated animals. In control patients, salivary gland function was significantly (p < 0.001) reduced by 40.2 +/- 14.1% and 39.9 +/- 15.3% in parotid and submandibular glands, respectively, three months after HD-RIT, and 11 patients developed xerostomia. In 25 amifostine-treated patients, salivary gland function was not significantly reduced (p = 0.691), and xerostomia did not occur. Thus, parenchymal damage in salivary glands induced by high-dose radioiodine therapy can be reduced significantly by amifostine. This may improve quality of life of patients with differentiated thyroid cancer.

Adult↗

Lymphoscintigraphy in tumors of the head and neck using double tracer technique.

UNLABELLED: Knowledge of possible lymphatic drainage may facilitate planning of surgery for patients with head and neck tumors. Therefore, the aim of this study was to present a method of lymphoscintigraphy with special attention to an accurate correlation of lymphatic drainage to anatomic regions. METHODS: Lymphoscintigraphy was performed using a double tracer technique before surgery in a total of 75 patients with squamous cell carcinoma of the head and neck. All patients received 100 MBq 99mTc-colloid at three to four peritumoral sites. A perchlorate solution (2 mL) was given orally to block salivary glands and the thyroid gland. Patients received 50 MBq 99mTc-pertechnetate intravenously for body contouring 20 min postinjection. Planar images were obtained over 5 min each, at 30 min and 4 h postinjection from anterior, right lateral and left lateral views with a large-field-of-view gamma camera. Lymphatic drainage was assessed by visual inspection and assigned to six cervical compartments. RESULTS: Neither the salivary glands nor the thyroid gland were seen in any of the patients. In 22 of 75 patients (29.3%), the injection site was the only focal tracer uptake seen. In contrast, lymphatic drainage was identified in the remaining 53 patients (70.7%), and lymph nodes could be assigned easily to the six cervical compartments. Of 75 patients, 36 (48%) exhibited ipsilateral lymphatic drainage. In addition, 17 patients (22.7%) with unilateral tumor showed bilateral (n = 12), contralateral (n = 2) or retropharyngeal (n = 3) lymphatic drainage. In 3 of these 17 patients, bilateral lymph node metastases were proven. A subgroup of 12 patients (16%) exhibited N2c nodal status, despite a unilateral localized primary tumor. In 3 of these 12 patients, surgery was extended as a result of scintigraphic findings from unilateral toward bilateral neck dissection, and histology confirmed nodal involvement in these patients. CONCLUSION: Lymphoscintigraphy using the double tracer technique allows an accurate correlation of lymphatic drainage to the six cervical compartments. This may provide the basis for a re-evaluation of its impact in treatment planning of patients with head and neck tumors.

Adult↗

Quantitative salivary gland scintigraphy.

OBJECTIVE: Uptake of 99mTc-pertechnetate in salivary glands reflects intact salivary gland parenchyma. However, no standardized protocol for an accurate quantification of parenchymal function has been established so far. METHODS: In this paper we report on a validated acquisition protocol supplying a normal database for standardized quantitative salivary gland scintigraphy. RESULTS: The major advantage of salivary gland scintigraphy, as compared to other imaging modalities, is that both parenchymal function and excretion fraction of all four major salivary glands (i.e., parotid and submandibular glands) can be simultaneously quantified with a single intravenous injection. CONCLUSION: Quantitative salivary gland scintigraphy is demonstrated to be a suitable imaging modality for research applications in evaluating the effects of radioprotective drugs on salivary glands. Salivary gland scintigraphy is easy to perform, reproducible and well-tolerated by the patient.

Adolescent↗