Growth Pattern in Chinese Children With 5α-Reductase Type 2 Deficiency: A Retrospective Multicenter Study

Background5α-reductase type 2 deficiency (5αRD) is an autosomal recessive hereditary disease of the group of 46, XY disorders of sex development (DSD).ObjectiveTo study the growth pattern in Chinese pediatric patients with 5αRD.SubjectsData were obtained from 141 patients with 5αRD (age: 0–16 years old) who visited eight pediatric endocrine centers from January 2010 to December 2017.MethodsIn this retrospective cohort study, height, weight, and other relevant data were collected from the multicenter hospital registration database. Baseline luteinizing hormone (LH), follicle stimulating hormone (FSH), testosterone (T), and dihydrotestosterone (DHT) after human chorionic gonadotropin (HCG) stimulation test were measured by enzyme enhanced chemiluminescence assay. Bone age (BA) was assessed using the Greulich-Pyle (G-P) atlas. Growth curve was constructed based on λ-median-coefficient of variation method (LMS).ResultsThe height standard deviation scores (HtSDS) and weight standard deviation scores (WtSDS) in 5αRD children were in the normal range as compared to normal boys. Significantly higher HtSDS was observed in patients with 5αRD who were <1 year old (t = 3.658, 2.103, P = 0.002, 0.048, respectively), and higher WtSDS in those <6 months old (t = 2.756, P = 0.012). Then HtSDS and WtSDS decreased gradually and fluctuated near the median of the same age until 13 years. WtSDS in 5αRD children from northern China were significantly higher than those from the south (Z = -2.670, P = 0.008). The variation tendency of HtSDS in Chinese 5αRDs was consistent with the trend of stimulating T. HtSDS and stimulating T in the external masculinization score (EMS) <7 group were slightly higher than those in EMS ≥ 7 group without significant difference. Additionally, the ratio of BA over chronological age (BA/CA) was significantly <1 in children with 5αRD.ConclusionChildren with 5αRD had a special growth pattern that was affected by high levels of T, while DHT played a very small role in it. Their growth accelerated at age <1 year, followed by slowing growth and fluctuating height near normal median boys’ height. The BA was delayed in 5αRD children. Androgen treatment, which may be considered anyway for male 5αRD patients with a micropenis, may also be beneficial for growth

Iron Uptake Mediated by Binding of H-Ferritin to the TIM-2 Receptor in Mouse Cells

Ferritin binds specifically and saturably to a variety of cell types, and recently several ferritin receptors have been cloned. TIM-2 is a specific receptor for H ferritin (HFt) in the mouse. TIM-2 is a member of the T cell immunoglobulin and mucin domain containing (TIM) protein family and plays an important role in immunity. The expression of TIM-2 outside of the immune system indicates that this receptor may have broader roles. We tested whether ferritin binding to TIM-2 can serve as an iron delivery mechanism. TIM-2 was transfected into normal (TCMK-1) mouse kidney cells, where it was appropriately expressed on the cell surface. HFt was labeled with (55)Fe and (55)Fe-HFt was incubated with TIM-2 positive cells or controls. (55)Fe-HFt uptake was observed only in TIM-2 positive cells. HFt uptake was also seen in A20 B cells, which express endogenous TIM-2. TIM-2 levels were not increased by iron chelation. Uptake of (55)Fe-HFt was specific and temperature-dependent. HFt taken up by TIM-2 positive cells transited through the endosome and eventually entered a lysosomal compartment, distinguishing the HFt pathway from that of transferrin, the classical vehicle for cellular iron delivery. Iron delivered following binding of HFt to TIM-2 entered the cytosol and became metabolically available, resulting in increased levels of endogenous intracellular ferritin. We conclude that TIM-2 can function as an iron uptake pathway

Iron uptake mediated by binding of H-ferritin to the TIM-2 receptor in mouse cells.

Ferritin binds specifically and saturably to a variety of cell types, and recently several ferritin receptors have been cloned. TIM-2 is a specific receptor for H ferritin (HFt) in the mouse. TIM-2 is a member of the T cell immunoglobulin and mucin domain containing (TIM) protein family and plays an important role in immunity. The expression of TIM-2 outside of the immune system indicates that this receptor may have broader roles. We tested whether ferritin binding to TIM-2 can serve as an iron delivery mechanism. TIM-2 was transfected into normal (TCMK-1) mouse kidney cells, where it was appropriately expressed on the cell surface. HFt was labeled with (55)Fe and (55)Fe-HFt was incubated with TIM-2 positive cells or controls. (55)Fe-HFt uptake was observed only in TIM-2 positive cells. HFt uptake was also seen in A20 B cells, which express endogenous TIM-2. TIM-2 levels were not increased by iron chelation. Uptake of (55)Fe-HFt was specific and temperature-dependent. HFt taken up by TIM-2 positive cells transited through the endosome and eventually entered a lysosomal compartment, distinguishing the HFt pathway from that of transferrin, the classical vehicle for cellular iron delivery. Iron delivered following binding of HFt to TIM-2 entered the cytosol and became metabolically available, resulting in increased levels of endogenous intracellular ferritin. We conclude that TIM-2 can function as an iron uptake pathway

Characterization of Tetratricopeptide Repeat-Containing Proteins Critical for Cilia Formation and Function

<div><p>Cilia formation and function require a special set of trafficking machinery termed intraflagellar transport (IFT), consisting mainly of protein complexes IFT-A, IFT-B, BBSome, and microtubule-dependent molecular motors. <u>T</u>etra<u>t</u>ri<u>c</u>opeptide repeat-containing (TTC) proteins are widely involved in protein complex formation. Nine of them are known to serve as components of the IFT or BBSome complexes. How many TTC proteins are cilia-related and how they function, however, remain unclear. Here we show that twenty TTC genes were upregulated by at least 2-fold during the differentiation of cultured mouse tracheal epithelial cells (MTECs) into multiciliated cells. Our systematic screen in zebrafish identified four novel TTC genes, <i>ttc4</i>, <i>-9c</i>, <i>-36</i>, and <i>-39c</i>, that are critical for cilia formation and motility. Accordingly, their zebrafish morphants displayed typical ciliopathy-related phenotypes, including curved body, abnormal otolith, hydrocephalus, and defective left-right patterning. The morphants of <i>ttc4</i> and <i>ttc25</i>, a known cilia-related gene, additionally showed pronephric cyst formation. Immunoprecipitation indicated associations of TTC4, -9c, -25, -36, and -39c with components or entire complexes of IFT-A, IFT-B, or BBSome, implying their participations in IFT or IFT-related activities. Our results provide a global view for the relationship between TTC proteins and cilia.</p></div

Screen for cilia-related TTC genes in zebrafish.

<p>(A) The morphologies of zebrafish morphants at 72 hpf. Note that the morphants of <i>ttc4</i>, <i>-9c</i>, <i>-25</i>, <i>-36</i>, and <i>-39c</i> displayed severe body curving. (B) Quantification results for the curved body phenotype. Three independent experiments were performed. (C) and (D) Rescue experiments for the <i>ttc</i> morphants. The indicated MO was coinjected with either the corresponding MO-resistant mRNA or GFP mRNA into zebrafish embryos. The body curvature phenotype was assayed at 72 hpf. The numbers of embryos counted are listed over the histograms. Statistical results are from three independent experiments. Student’s <i>t</i>-test, ***P < 0.001. Error bars represent s.d.</p

The expression profiles of TTC genes upregulated during MTEC differentiation into multiciliated cells.

<p>(A) Typical differentiation progression of MTECs cultured <i>in vitro</i>. MTECs were isolated from mice of 8-week old and were induced to differentiate into multiciliated cells by culturing at an air-liquid interface (ALI) for the indicated time. Approximately 90% of MTECs were multiciliated at ALI day 5. Acetylated tubulin (Ace-tub) was used to mark cilia. ZO-1 labels the tight junctions. (B) The gene expression profiles of known cilia-related TPR-containing proteins following the MTEC differentiation. The data were from cDNA microarray analyses on MTECs samples from ALI d0 to ALI d5. The gene expression pattern of Deup1, a protein critical for centriole amplification, is also included for comparison. (C) The expression profiles of ten novel TTC genes. (D) Expression profiles of the indicated genes, based on quantitative PCR (qPCR) results. One set of typical results of two independent experiments is presented (Please see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0124378#pone.0124378.s002" target="_blank">S2 Fig</a> for the second set of results). The mRNA levels at ALI d0 were set as 1.</p

Examinations on cilia in the Kupffer’s vesicle and pronephric duct.

<p>(A-C) Both cilia number and cilia length in the Kupffer’s vesicle (KV) at the 7-somite stage were reduced in the indicated <i>ttc</i> morphants. Cilia were labeled using antibody against acetylated tubulin. The quantification results were based on three independent experiments. Student’s <i>t</i>-test, ***P < 0.001. Error bars represent s.d. (D) Typical morphology of cilia in pronephric ducts at 24 hpf. (E) and (F) Cilia motility in the pronephric duct at 60 hpf. The kymographs showed trajectories of the cilia marked with red line in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0124378#pone.0124378.s010" target="_blank">S1</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0124378#pone.0124378.s011" target="_blank">S2</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0124378#pone.0124378.s012" target="_blank">S3</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0124378#pone.0124378.s013" target="_blank">S4</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0124378#pone.0124378.s014" target="_blank">S5</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0124378#pone.0124378.s015" target="_blank">S6</a> Videos. Cilia beat frequencies (CBFs) were measured at 60 hpf (40 cilia from 4 morphants for each gene). Student’s <i>t</i>-test, ***P < 0.001. Error bars represent s.e.m.</p