Biomedical subjects

G Schmitz

Publications and source records attributed to G Schmitz.

At least 37 records · Page 2Linked to original sources

PubMed2002

Identification of regulatory elements in the human adipose most abundant gene transcript-1 ( apM-1) promoter: role of SP1/SP3 and TNF-alpha as regulatory pathways.

AIMS/HYPOTHESIS: The human adipocyte-specific apM-1 (adipose most abundant gene transcript-1) gene encodes for a secretory protein of the adipose tissue that seems to play a role in the pathogenesis of obesity-related insulin resistance and its expression is inhibited by TNF-alpha. Our aim was to characterize the tissue-specific regulation of the recently cloned apM-1 promoter and the mechanisms of TNF-alpha-induced downregulation of the apM-1 gene. METHODS: We characterised the apM-1 gene by electrophoretic mobility shift assays (EMSA) and luciferase reporter gene assays (LRA). RESULTS: Although several putative binding sites for transcription factors known to be involved in adipogenesis such as C/EBP and PPARgamma are present in the promoter, we could not detect any binding of these nuclear proteins from differentiated adipocytes. However, a proximal SP1 binding site specifically binds both, recombinant SP1 protein and SP1 derived from adipocyte nuclear extracts. Since the expression of SP1 during adipocyte differentiation has not yet been analysed, we could show by using EMSA, that binding activity of SP1 is increased during adipocyte differentiation. The stimulatory activity of SP1 was confirmed in LRA by cotransfection experiments in S2 Schneider cells lacking endogenous SP factors. An inhibitory activity of SP3 on the stimulatory effect of SP1 could be confirmed in LRA by contransfection experiments in adipocytes. Nuclear extracts from adipocytes incubated with TNF-alpha showed a reduced binding activity of SP1. CONCLUSION/INTERPRETATION: SP1 is expressed and its binding activity is enhanced during adipocyte differentiation. SP1 has stimulatory effects, SP3 has inhibitory effects on apM-1 promoter activity, mediated by a proximal SP1 binding site. The mechanism of TNF-alpha-induced inhibition of apM-1 gene expression is, at least in part, due to a decrease of transcriptional SP1 binding activity caused by TNF-alpha and thus provides a new mechanism of TNF-alpha-dependent signalling.

3T3 Cells↗

PubMed2002

ABCA2: a candidate regulator of neural transmembrane lipid transport.

Studies in the past years have implicated multispan transmembrane transport molecules of the ATP binding cassette (ABC) transporter family in cellular lipid export processes. The prototypic ABC transporter ABCA1 has recently been demonstrated to act as a major facilitator of cellular cholesterol and phospholipid export. Moreover, the transporter ABCA4 (ABCR) plays a pivotal role in retinaldehyde processing, and ABCA3 has recently implicated in lung surfactant processing. These pioneering observations have directed considerable attention to the A subfamily of ABC proteins. ABCA2 is the codefining member of the ABC A-transporter subclass. Although known for some time, it was not until recently that its complete molecular structure was established. Unlike other ABC A-subfamily members, ABCA2 is predominantly expressed in the brain and neural tissues. The unique expression profile together with available structural data suggest roles for this largest known ABC protein in neural transmembrane lipid export.

ATP-Binding Cassette Transporters↗

PubMed2002

Thiopental impairs neutrophil oxidative response by inhibition of intracellular signalling.

BACKGROUND AND OBJECTIVE: Thiopental in clinically relevant concentrations inhibits the oxidative function of neutrophils, whereas only very high, non-therapeutic concentrations of methohexital induce a similar effect. The study characterized the molecular basis of this differential action of oxy- and thiobarbiturates on neutrophils. METHODS: Neutrophils were incubated in vitro with thiopental or methohexital using concentrations within the therapeutic range. Neutrophil responses were induced using different stimuli: N-formyl-L-methionyl-L-leucyl-L-phenylalanine (FMLP), C5a and 1,2-dioctanoyl-sn-glycerol (DiC8-DAG). FMLP and C5a bind to specific G-protein-coupled receptors that share the same second messenger cascade. In contrast, DiC8-DAG, an activator of protein kinase C, bypasses the signal transduction pathway downstream of the receptors. Hydrogen peroxide production by neutrophils was assessed using flow cytometry. To characterize the localization of the interaction site, FMLP receptor expression and cytosolic-free calcium were further analysed. RESULTS: FMLP and C5a-induced hydrogen peroxide production were both significantly impaired by thiopental, but not by methohexital. When postreceptor signalling was bypassed, by stimulation with DiC8-DAG, neither thiopental nor methohexital affected hydrogen peroxide production. Additionally, neither of the barbiturates impaired the cytosolic Ca2+ response. CONCLUSIONS: We conclude that neither protein kinase C nor the hydrogen peroxide-generating enzymes are affected by thiopental or methohexital. The unimpaired Ca2+ response suggests that the function of the receptors and G-proteins were also unimpaired. Taken together, this indicates that the site of action of thiopental is in the cellular signalling upstream of protein kinase C.

Anesthetics, Intravenous↗

PubMed2002

Identification of novel mutations in the NPC1 gene in German patients with Niemann-Pick C disease.

Niemann-Pick disease type C (NPC) is an inherited neuro-degenerative disorder associated with intracellular cholesterol trafficking defects. Mutations in two distinct genes, NPC1 and HE1, have recently been shown to cause this disease. We have analysed the NPC1 gene in five German patients with NPC from four unrelated families. We identified a total of five novel mutations in the coding region of the NPC1 gene (G231V, D874V, 1642M, 11094T and R116stop). All affected individuals displayed compound heterozygosity. The mutated alleles were transmitted by the nonaffected parents with the exception of one patient, in whom a de novo mutation (G231V) had occurred. Interestingly, the G231V/P237S NPC1 genotype in this individual is associated with an early-onset form of NPC. In contrast, we found that the D874V/D948N genotype, observed in another NPC patient, is characterized by a late onset of clinical symptoms that presents with a pronounced white-matter disease. Our results will contribute to defining the association between the clinical phenotypes and the genetic abnormalities in Niemann-Pick C disease.

Adolescent↗

PubMed2002

LDL-oxidation and -subgroups in normo- and hypertensive patients with and without a family history of hypertension.

OBJECTIVES: Oxidative modification of low-density-lipoprotein (LDL) increases its atherogenic potential to induce the accumulation of lipids and cells in the vascular wall. Patients have different lipoprotein profiles according to their LDL-subgroup pattern. The subgroup of LDL, which is most susceptible to oxidation, is most likely the dense LDL3 subfraction. In order to study an assumed association between hypertension, LDL subgroup distribution and the susceptibility of LDL to oxidation, 14 normotensive patients without family histories of hypertension (NT), 13 normotensive patients with family histories of hypertension (NT-FH), 10 hypertensive patients without family histories of (HT) and 11 hypertensive patients with family histories of hypertension (HT-FH) were evaluated. PATIENTS AND METHODS: LDL was oxidatively modified by incubation with copper ions (1.6 microM/L). The course of LDL-oxidation was measured in vitro by continuous photometric monitoring and the quantitative distribution of 3 LDL-subgroups by capillary isotachophoresis (ITP). RESULTS: The lag-phases of NT-FH and hypertensive patients were shorter than those of the control group (NT: 116 +/- 36 minutes; NT-FH 92 +/- 32 minutes, p < 0.05; HT: 95 +/- 41 minutes; HT-FH: 76 +/- 33 minutes, p < 0.05). Compared to NT a significant difference in the relative preponderance of LDL3 subgroup was observed for HT-FH (23.5 +/- 4.6% versus NT: 19.3 +/- 6.6%), additionally, statistical analysis showed a similar trend amongst the other patient groups (NT-FH: 20.4 +/- 7.4%, HT: 21.4 +/- 4.6%). CONCLUSIONS: The increased occurrence of the LDL3 subgroup might contribute to a higher susceptibility to LDL oxidation and therefore create an increased risk of vascular disease in the genotypic and phenotypic hypertensive patient population.

Adult↗

PubMed2002

HLA-B*3531, a hybrid of B35 and B61, implications for diagnostic approaches to alleles with complex ancestral compositions.

The serological characterization of allelic variants that have been generated by large-scale interallelic recombination events indicates which residues may be involved in the formation of epitopes crucial for serological recognition. The allelic product of HLA-B*3531 is composed of B35 in its alpha1 domain and of B61(40) in its alpha2 domain. Both specificities are only weakly detectable with available sera. Allelic products with 'mixed' serology also represent a challenge to DNA-based HLA typing methods, as only the sequence motif of one ancestral allele may be recognized. In this case the hidden specificity would not be considered in the matching process and might not be recognized as an antigen 'unacceptable' to the recipient.

Alleles↗

PubMed2002

Pro12Ala polymorphism in the peroxisome proliferator-activated receptor-gamma2 (PPARgamma2) is associated with higher levels of total cholesterol and LDL-cholesterol in male caucasian type 2 diabetes patients.

UNLABELLED: The peroxisome proliferator-activated receptor-gamma2 (PPAR(gamma2)) represents the transcriptional master regulator of adipocyte differentiation and therefore has been suggested as candidate gene for the pathogenesis of obesity, type 2 diabetes and related metabolic disorders. Aim of our study was to determine the frequency of a missense point mutation within exon 2 of PPAR(gamma2), Pro12Ala, and its possible association with metabolic parameters as well as diabetic retinopathy (in a population-based sample of 560 (318 male ad 242 female) type 2 diabetic patients. Subsequent to genomic PCR amplification, the Hpa-II RFLP analysis was used for genotyping. RESULTS: 436 (77.9%) subjects were homozygous for the wildtype allele (Pro/Pro), 118 (21.1%) were heterozygous (Pro/Ala) and 6 (1.1%) were homozygous for the mutated allele (Ala/Ala). Genotype frequency was calculated to be 0.81 for the wildtype and 0.19 for the mutated allele. These frequencies did not differ from non-diabetic cohorts examined earlier. In contrast to females, total cholesterol and LDL-cholesterol were significantly higher in males (Total cholesterol: 281.8 +/- 51.3 vs 253.1 +/- 49.8 mg/dl, p < 0.0001; LDL-cholesterol: 182.0 +/- 49.2 vs 155.6 +/- 42.0 mg/d, p < 0.0001) in the presence of the mutated allele as compared to the wildtype subgroup. No differences were found with respect to BMI, HbA1c, blood pressure and serum levels of leptin nor to prevalence of retinopathy. Pro12Ala polymorphism of PPAR(gamma2) gene is not associated with diabetic retinopathy but is associated with dyslipidemia in male type 2 diabetic patients.

Aged↗

PubMed2002

Definition and characterization of relative hypo- and hyperleptinemia in a large Caucasian population.

The adipocyte-derived hormone, leptin, has been implicated in the regulation of appetite, weight gain and glucose homeostasis as well as in liver fibrogenesis, hematopoiesis and immune function. No previous reports have clearly defined pathologically elevated or decreased serum leptin levels for Caucasian adults. The aim of this study was to define and characterize subjects with relative hyper- and hypoleptinemia in a large population-based German cohort. Percentiles of leptin levels by body mass index (BMI) were calculated from 4971 adult Germans, and the participants with leptin levels above the 95th and below the 5th percentile were defined as relatively hyperleptinemic and relatively hypoleptinemic, respectively, for their BMI. These participants were compared with the intermediate group with respect to anthropometric and clinical data and parameters of glucose and iron metabolism, lipid status, renal, adrenal and reproductive function. Relatively hyperleptinemic participants (HL) showed higher insulin, c-peptide, and total cholesterol levels than the hypoleptinemic subjects; in males, ferritin levels were higher and testosterone levels lower in the HL group. In conclusion, we report the first percentile curves for serum leptin by BMI in a large Caucasian population. Relatively low leptin values may be associated with a lower metabolic risk than relatively high serum leptin values.

Adult↗

PubMed2001

Characterization of the ATPase cycle of human ABCA1: implications for its function as a regulator rather than an active transporter.

ABCA1 plays a key role in cellular cholesterol and phospholipid traffic. To explore the biochemical properties of this membrane protein we applied a Baculovirus-insect cell expression system. We found that human ABCA1 in isolated membranes showed a specific, Mg(2+)-dependent ATP binding but had no measurable ATPase activity. Nevertheless, conformational changes in ABCA1 could be demonstrated by nucleotide occlusion, even without arresting the catalytic cycle by phosphate-mimicking anions. Addition of potential lipid substrates or lipid acceptors (apolipoprotein A-I) did not modify the ATPase activity or nucleotide occlusion by ABCA1. Our data indicate that ATP hydrolysis by ABCA1 occurs at a very low rate, suggesting that ABCA1 may not function as an effective active transporter as previously assumed. In the light of the observed conformational changes we propose a regulatory function for human ABCA1.

ATP Binding Cassette Transporter 1↗

PubMed2001

The role of plasma phospholipid transfer protein (PLTP) in HDL remodeling in acute-phase patients.

During reverse cholesterol transport plasma phospholipid transfer protein (PLTP) converts high density lipoprotein(3) (HDL(3)) into two new subpopulations, HDL(2)-like particles and pre-beta-HDL. The acute-phase response is accompanied with dramatic changes in lipid metabolism including alterations in HDL concentration, composition, and thereby its function as a substrate for HDL remodeling proteins in circulation. To evaluate how acute-phase HDL (AP-HDL) functions in PLTP-mediated HDL conversion, we collected plasma samples from patients with severe acute-phase response (n=17), and from healthy controls (n=30). Subsequently, total HDL (1.063<d<1.21 g/ml) was isolated from patients and controls and incubated in the absence and presence of purified PLTP. The results show that HDL isolated from the acute-phase patients is converted by PLTP in vitro in a corresponding manner as normal HDL. In the combined population, C-reactive protein correlated significantly with lecithin-cholesterol acyltransferase (LCAT) activity (r=-0.53), cholesterol ester transfer protein activity (r=-0.80), PLTP activity (r=0.44), and PLTP mass (r=-0.66). When compared to the controls, the patients had 31% higher PLTP activity, but 52% lower PLTP mass leading to a 165% higher PLTP specific activity in the patients. The present data indicate that during the acute-phase response, plasma PLTP activity and mass are strongly affected by the lipoprotein distribution as well as lipid composition. We suggest that the decrease of HDL during the acute phase is caused by reduced LCAT and increased PLTP activities both increasing the plasma levels of lipid-poor apoA-I particles.

Acute-Phase Reaction↗

PubMed2001

ABCA6, a novel a subclass ABC transporter.

Here we report the cDNA cloning of a novel member of the ABC A transporter subfamily from human macrophages. The identified coding sequence is of 5.0 kb size and contains an open reading frame which encodes a 1617 amino acid polypeptide. Structurally, the putative ABC transporter protein product consists of two tandemly oriented subunits, each composed of a transmembrane domain followed by a nucleotide binding fold, and thus conforms to the group of full-size ABC transporters. We also demonstrate the existence of an alternative transcript that codes for a 637 amino acid protein variant bearing the features of a truncated half-size transporter. Among the human ABC transporter subfamily A the novel transporter shows highest protein sequence homology with ABCA8 (60%), followed by ABCA2 (32%) and ABCA1 (32%), respectively. In agreement with the proposed classification, the novel transporter was designated ABCA6. The ABCA6 gene is ubiquitously expressed with highest mRNA levels in liver, lung, heart and brain. Analysis of the genomic organization demonstrated that the ABCA6 gene is composed of 38 exons which extend across a region of 62 kb size on chromosome 17q24.2. Based on its structural features and its cholesterol-responsive regulation ABCA6 is potentially involved in macrophage lipid homeostasis.

ATP Binding Cassette Transporter 1↗

PubMed2001

Homogeneous assay based on 52 primer sets to scan for mutations of the ABCA1 gene and its application in genetic analysis of a new patient with familial high-density lipoprotein deficiency syndrome.

Familial high-density lipoprotein (HDL)-deficiency syndromes are caused by mutations of the ABCA1 gene, coding for the ATP-binding cassette transporter 1. We have developed a homogeneous assay based on 52 primer sets to amplify all 50 ABCA1 exons and approximately 1 kb of its promoter. The assay allows for convenient amplification of the gene from genomic DNA and easy mutational analysis through automatic sequencing. It obviates the need to use mRNA preparations, which were difficult to handle and posed a risk to miss splice junction or promoter mutations. The application of the test to the molecular analysis of a new patient with familial HDL-deficiency (Tangier disease) led to a discovery of two novel ABCA1 mutations: C2665del and C4457T.

ATP Binding Cassette Transporter 1↗

PubMed2001

Adipophilin is a sensitive marker for lipid loading in human blood monocytes.

Adipophilin, a marker of lipid accumulation initially described in adipocytes, was recently shown to be induced in macrophage foam cells. We found that even freshly isolated blood monocytes express adipophilin and that the amount of adipophilin protein is variable in monocytes from different healthy individuals. However, the physiological expression of adipophilin does not correlate with the levels of free fatty acids, cholesterylesters or free cholesterol. Enzymatically modified low-density lipoprotein (E-LDL) induces rapid foam cell formation in monocytes and upregulates adipophilin mRNA and protein within 2 h of incubation. This rapid induction of adipophilin is accompanied by a significant increase of free fatty acids in monocytes incubated with E-LDL. Adipophilin facilitates the uptake of free fatty acids, and here we demonstrate that free fatty acids increase is related to the early upregulation of adipophilin expression in blood monocytes. Fatty acids are ligands for peroxisome proliferator-activated receptor-gamma (PPARgamma), and the upregulation of adipophilin mRNA by PPARgamma agonists like 15d-PGJ(2) and ciglitazone indicates that PPARgamma may mediate the induction of adipophilin expression in human blood monocytes.

Adult↗

PubMed2001

Inhibition of effects of endogenously synthesized histamine disturbs in vitro human dendritic cell differentiation.

Histamine, a principal mediator in various physiological and pathological cell functions is synthesized from L-histidine exclusively by histidine decarboxylase, an enzyme, which is expressed in many tissues of mammalian organism. Histamine plays a role in various cellular functions, including cell differentiation. The aim of this study was to determine the presence and to characterize the role of the endogenously produced histamine during in vitro dendritic cell (DC) differentiation induced by interleukin-4 (IL-4) and granulocyte-monocyte colony stimulating factor (GM-CSF). The changes in intracellular histamine content, biosynthesis and gene expression of histidine decarboxylase were investigated during this process. One also studied how histamine receptor antagonists and a histamine synthesis blocker influence the expression of differentiation antigens on the DC during in vitro maturation. During in vitro differentiation parallel culture incubations were performed by adding H1 receptor antagonist triprolidine, H2 receptor antagonist tiotidine, the tamoxifene derivate DPPE which blocks the intracellular binding of histamine, and an irreversible blocker of histidine decarboxylase, alpha-fluoromethyl histamine (alpha-FMH). The results show simultaneous increase in both histidine decarboxylase level and histamine content during differentiation of elutriated monocytes toward DC. Both blockade of de novo histamine production (by alpha-FMH) and inhibition of histamine binding (by H1 and H2 receptor antagonists, triprolidine and tiotidine, respectively) markedly decreased CD40 expression and that of CD45 from the 3rd day of treatment. DPPE by disturbing intracellular interaction of histamine with cytochrome P-450 moieties was able to decrease the expression of CD45, CD86, HLA-DR, CD33, CD40 and CD11c. Based on the data it is suggested that endogenous histamine is actively synthesized during cytokine-induced in vitro DC differentiation. The functional relevance and autocrine and paracrine action of endogenously produced histamine is supported by the data showing that inhibition of histamine synthesis by HDC, blocking of histamine binding by both 'extracellular' histamine receptors (by specific antagonists, triprolidine and tiotidine) and 'intracellular' antagonists (DPPE) disturb the differentiation of DC. This conclusion is supported by the fact, that by the inhibition of histamine acting in an autocrine/paracrine way, the expression pattern of differentiation markers on DC is markedly changed.

Antigens, CD↗

PubMed2001

ABC transporters and cholesterol metabolism.

ATP-binding cassette (ABC) proteins form a group of highly conserved cellular transmembrane transporters. Studies over the past year have implicated ABC transporters in cellular lipid trafficking processes. This notion has recently been confirmed and extended by the finding that the ABC transporter ABCA1 is a key regulator of high-density lipoprotein (HDL) metabolism and macrophage targeting to the RES or the vascular wall. Expression of a large number of ABC transporters in monocytes/macrophages and their regulation by cholesterol flux render these transporter molecules potentially critical players in chronic inflammatory diseases such as atherosclerosis.

ATP-Binding Cassette Transporters↗

PubMed2001

Complete coding sequence, promoter region, and genomic structure of the human ABCA2 gene and evidence for sterol-dependent regulation in macrophages.

Members of the human ABC transporter A subfamily have gained considerable attention based on the recent findings that ABCA1 and ABCR (ABCA4) cause familial HDL-deficiency syndromes and distinct forms of hereditary retinopathies, respectively. Here we report the complete cDNA and the genomic organization of ABCA2, another member of the human ABC A transporter subfamily. The ABCA2 coding region is 7.3 kb in size and codes for a 2436 amino acid polypeptide that bears the typical features of a full-size ABC transporter. Among the known members of the ABC A subfamily ABCA2 shares highest homology with the cholesterol-responsive transporters ABCA1 (50%) and the recently cloned ABCA7 (44%). The ABCA2 gene comprises 48 exons which are localized within a genomic region of only 21 kb. Analysis of the putative ABCA2 promoter sequence revealed potential binding sites for transcription factors that are involved in the differentiation of myeloid and neural cells. Gene expression analysis in human macrophages showed that ABCA2 mRNA is induced during cholesterol import indicating that ABCA2 is a cholesterol-responsive gene. Our results suggest a potential role for ABCA2 in macrophage lipid metabolism and neural development.

ATP-Binding Cassette Transporters↗

PubMed2001

Common variants in the gene encoding ATP-binding cassette transporter 1 in men with low HDL cholesterol levels and coronary heart disease.

HDL cholesterol (HDL-C) deficiency is the most common lipid abnormality observed in patients with premature coronary heart disease (CHD). Recently, our laboratory and others demonstrated that mutations in the ATP-binding cassette transporter 1 (ABCA1) gene are responsible for Tangier disease, a rare genetic disorder characterized by severely diminished plasma HDL-C concentrations and a predisposition for CHD. To address the question of whether common variants within the coding sequence of ABCA1 may affect plasma HDL-C levels and CHD risk in the general population, we determined the frequencies of three common ABCA1 variants (G596A, A2589G and G3456C) in men participating in the Veterans Affairs Cooperative HDL Cholesterol Intervention Trial (VA-HIT), a study designed to examine the benefits of HDL raising in men having low HDL-C (< or =40 mg/dl) and established CHD, as well as in CHD-free men from the Framingham Offspring Study (FOS). Allele frequencies (%) in VA-HIT were 31, 16, and 4 for the G596A, A2589G, and G3456C variants, respectively, versus 27, 12, and 2 in FOS (P<0.03). None of the variants were significantly associated with plasma HDL-C concentrations in either population; however, in VA-HIT, the G3456C variant was associated with a significantly increased risk for CHD end points, suggesting a role for this variant in the premature CHD observed in this population.

ATP Binding Cassette Transporter 1↗

PubMed2001

Lipoprotein (a) up-regulates the expression of the plasminogen activator inhibitor 2 in human blood monocytes.

Elevated plasma lipoprotein (a) (Lp[a]) and cardiac events show a modest but significant association in various clinical studies. However, the influence of high Lp(a) on the gene expression in blood monocytes as a major cell involved in atherogenesis is poorly described. To identify genes influenced by elevated serum Lp(a), the gene expression was analyzed on a complementary DNA microarray comparing monocytes from a patient with isolated Lp(a) hyperlipidemia and coronary heart disease with monocytes from a healthy blood donor with low Lp(a). By using this approach, numerous genes were found differentially expressed in patient-versus-control monocytes. Verification of these candidates by Northern blot analysis or semiquantitative polymerase chain reaction in monocytes from additional patients with Lp(a) hyperlipidemia and healthy blood donors with elevated Lp(a) confirmed a significant induction of plasminogen activator inhibitor type 2 (PAI-2) messenger RNA (mRNA) in monocytes from male, but not from female, individuals with high Lp(a), indicating that this observation is gender specific. This led also to increased intracellular and secreted PAI-2 protein in monocytes from male probands with Lp(a) hyperlipidemia. Plasminogen activator inhibitor type 1 (PAI-1) mRNA was found suppressed only in the patients' monocytes and not in healthy probands with high Lp(a) levels. Purified Lp(a) induced PAI-2 mRNA and protein and reduced PAI-1 expression in monocytes isolated from various controls. The finding that PAI-2 is elevated in monocytes from male patients with isolated Lp(a) hyperlipidemia and male healthy probands with high Lp(a) and that purified Lp(a) up-regulates PAI-2 in control monocytes in vitro indicate a direct, but gender-specific, effect of Lp(a) for the induction of PAI-2 expression.

Adult↗