Spinal Stenosis and Thoracolumbar Kyphosis in Achondroplasia / 罕见病研究

Achondroplasia (ACH) is a rare autosomal-dominant genetic disease resulting from a mutation in the fibroblast growth factor receptor-3 (FGFR3) gene. It is characterized by asymmetric short stature. Spinal stenosis and thoracolumbar kyphosis (TLK) are common findings in ACH patients. Severe TLK can exacerbate spinal stenosis, leading to neurological complications. This paper provides a brief review of the pathophysiological mechanisms, clinical characteristics, and treatments for spinal stenosis and TLK in ACH patients. Recently, three new drugs targeting FGFR3; vosoritide, recifercept, and infigratinib, have completed or are undergoing clinical trials. They have shown promising preliminary results in preventing spinal stenosis and TLK.

Clinical features and FGFR3 mutations of children with achondroplasia / 中国当代儿科杂志

OBJECTIVES@#To study the clinical features and fibroblast growth factor receptor 3 (FGFR3) gene mutations of children with achondroplasia (ACH) through an analysis of 17 cases.@*METHODS@#A retrospective analysis was performed on the clinical data and FGFR3 gene detection results of 17 children with ACH who were diagnosed from January 2009 to October 2021.@*RESULTS@#Of the 17 children with ACH, common clinical manifestations included disproportionate short stature (100%, 17/17), macrocephaly (100%, 17/17), trident hand (82%, 14/17), and genu varum (88%, 15/17). The common imaging findings were rhizomelic shortening of the long bones (100%, 17/17) and narrowing of the lumbar intervertebral space (88%, 15/17). Major complications included skeletal dysplasia (100%, 17/17), middle ear dysfunction (82%, 14/17), motor/language developmental delay (88%, 15/17), chronic pain (59%, 10/17), sleep apnea (53%, 9/17), obesity (41%, 7/17), foramen magnum stenosis (35%, 6/17), and hydrocephalus (24%, 4/17). All 17 children (100%) had FGFR3 mutations, among whom 13 had c.1138G>A hotspot mutations of the FGFR3 gene, 2 had c.1138G>C mutations of the FGFR3 gene, and 2 had unreported mutations, with c.1252C>T mutations of the FGFR3 gene in one child and c.445+2_445+5delTAGG mutations of the FGFR3 gene in the other child.@*CONCLUSIONS@#This study identifies the unreported mutation sites of the FGFR3 gene, which extends the gene mutation spectrum of ACH. ACH is a progressive disease requiring lifelong management through multidisciplinary collaboration.

Achondroplasia: a case report and the review of the basics

The achondroplasia is a variant of short-limbed dwarfism. The word achondroplasia literally means without cartilage formation. However, in achondroplasia the problem is not in formation of cartilage but, in its conversion to bone (i.e. ossification). This deficient ossification is particularly seen in the long bones of arm and leg. The characteristic external appearance of people born with achondroplasia is short stature. The average height of an adult male with achondroplasia is 131 centimetres (4 feet, 4 inches), and the average height for adult females is 124 centimetres (4 feet, 1 inch). The trunk is of average size but the leg and upper arm is of short length. It is because the femur and humerus are relatively shorter in length. The range of movement at elbow is limited. The head is enlarged called macrocephaly and is with a prominent forehead. People with Achondroplasia are generally of normal intelligence. They have bowed legs and abnormal curvature of spine giving rise to lordosis or kyphosis. They may develop spinal stenosis, which is associated with pain, tingling and weakness in leg. This may cause difficulty in walking. The other health problems associated with Achondroplasia are episodes of apnoea, obesity and recurrent ear infection. The purpose of this study is to evaluate the cardinal phenotypic features in patient of Achondroplasia. It is also to assess the body physique, anthropometric measurements and to study the typical radiological signs in such patients as the main tool of diagnosis.

Expression of FGFR3 in acute lymphoblastic leukemia patients and its contribution to proliferation of circulating endothelial cells / 中国病理生理杂志

AIM: To evaluate the expression of fibroblast growth factor receptor 3 (FGFR3) in acute lymphocytic leukemia (ALL) patients and its contribution to the proliferation of circulating endothelial cells (CECs).METHODS: The mRNA expression levels of FGFR3 in 44 patients with ALL were assayed by RT-PCR.Overall survival (OS) rates of the patients in FGFR3+ group and FGFR3-group were estimated by Kaplan-Meier analysis.The CECs were sorted from peripheral blood by magnetic-activated cell sorting and then counted by 3-color flow cytometry.The cell counts, antigen expression, growth curve and colony forming rate of the CECs in the 2 groups were determined.The FGFR3 expression of CECs was identified by immunofluorescence staining.RESULTS: The positive rate of FGFR3 mRNA expression was 43.2% in 44 ALL patients with normal karyotype.T-ALL expressed higher level of FGFR3 than B-ALL (P<0.05).FGFR3 was over-expressed in ALL patients with bone marrow blast proportion ≥5% (P<0.05).The probability of OS was significantly lower in FGFR3+ group than that in FGFR3-group (P<0.05).The sorted CECs highly expressed CD31, CD144, VEGFR-2 and CD146, and rarely expressed CD45.The counts of CECs and expression level of CD133 significantly increased in FGFR3+ group compared with FGFR3-group.The same result of the amount of colony formation was observed (P<0.05).There was significant difference at 3 time points of cultured CECs count in vitro between FGFR3+ group and FGFR3-group (P<0.05).The positive rate of FGFR3 expression of CECs from 19 ALL-FGFR3+ patients was (29.00±15.71)%.CONCLUSION: The over-expression of FGFR3 gene in ALL may be helpful to evaluate the prolife-ration of CECs, and become a double target with anti-tumor and anti-angiogenesis effects to offer more choice for molecular therapy in the future.

miR-100 regulates cell growth through targeting FGFR3 in patients with pancreatic cancer / 中华肝胆外科杂志

Objective To investigate the effects of miR-100 on the proliferation of MIA PaCa-2 and CFPAC-1 cells through targeting fibroblast growth factor receptor 3 (FGFR3).Methods miR-100 expression levels in 17 cancer tissues and 17 nonmalignant tissues were examined by Real-time PCR.The effect of miR-100 overexpression on cell proliferation was examined by CCK-8 assay in vitro.Luciferase assay was used to confirm that miR-100 could directly target FGFR3.Real-time PCR and Western blot were used to examine the expression of FGFR3 in miR-100 overexpressing pancreatic cancer cells.The predicted target gene of miR-100,FGFR3,was downregulated by siRNA,and its effect on cell proliferation was also examined.Cell proliferation was analyzed using CCK-8 and Edu assay.Results miR-100 was lowly expressed in pancreatic cancer tissues (P ＜ 0.05).In pancreatic cancer cells,the transfection of lv-miR-100 was able to upregulate the endogenous expression of miR-100 and inhibit the cell proliferation (P ＜0.05).Luciferase assay showed FGFR3 was the direct target of miR-1O0.FGFR3 was significantly downregulated by overexpressing miR-100 in pancreatic cancer cells (P ＜0.05),and FGFR3 knockdown by specific siRNA exerted the similar effect as miR-100 overexpression (P ＜ 0.05).Conclusions Our study identified a new miRNA regulator,miR-100,and clarified a novel mechanism of how miR-100 regulates cell proliferation in pancreatic cancer.The strategy of overexpressing the tumor suppressor miR-100 may provide a new therapeutic approach for treating patients with pancreatic cancer.

Expression and purification of soluble single-chain Fv against human fibroblast growth factor receptor 3 fused with Sumo tag in Escherichia coli

Background Overexpression or mutated activation of Fibroblast growth factor receptor 3 (FGFR3) is involved in the pathogenesis of many tumors. More and more studies focus on the potential usage of therapeutic antibodies against FGFR3. Results In this study, a novel single-chain Fv (ScFv) against FGFR3 was prepared and characterized. To achieve the soluble expression, ScFv was fused with Sumo (Small ubiquitin-related modifier) by polymerase chain reaction (PCR), and cloned into pET-20b. The recombinant bacteria were induced by 0.5 mM Isopropyl-ß-d-thiogalactopyranoside (IPTG) for 16 h at 20°C, and the supernatant liquid of Sumo-ScFv was harvested and purified by Ni-NTA chromatography. After being cleaved by the Sumo protease, the recombinant ScFv was released from the fusion protein, and further purified by Ni-NTA chromatography. The purity of ScFv was shown to be higher than 95% and their yield reached 4 mg per liter of bacterial culture. In vitro data showed that ScFv can significantly attenuate FGF9-induced phosphorylation of FGFR3. Conclusion We provide a novel method to produce soluble expression and bioactive functions of ScFv in Escherichia coli.

Delayed Presentation of Rickets in a Child with Labyrinthine Aplasia, Microtia and Microdontia (LAMM) Syndrome.

Background: Labyrinthine Aplasia, Microtia and Microdontia (LAMM) syndrome is characterized by the complete absence of inner ear structures (Michel aplasia), microtia and microdontia. Hypophosphatemic rickets results from defects in the renal tubular reabsorption of filtered phosphate. Case characteristics: 13-year-old Indian girl presented with deafness since infancy and progressive wrist widening and genu valgum for last one year. Observation: Homozygous novel missense mutation in fibroblast growth factor 3. Message: LAMM syndrome and hypophosphatemic rickets may be associated.

Antenatal diagnosis of Thanatophoric dysplasia: a case report and review of literature.

Thanatophoric dysplasia is the lethal skeletal dysplasia characterized by marked underdevelopment of the skeleton and short-limb dwarfism. Foetus has short neck, narrow thoracic cage and protruberant abdomen. Other anatomical features include a relatively enlarged head with frontal bossing, platyspondyly, telephone handle femurs. Thanatophoric dysplasia is usually diagnosed using second trimester ultrasonography. We report this rare entity from rural India with emphasis on its anatomical features along with review of relevant literature.

FGFR3 gene mutation analysis of achondroplasia and hypochondroplasia families / 临床儿科杂志

Objective To screen the sequence of fibroblast growth factor receptor 3 (FGFR3) genes in children with dys-chondroplasia and their family members for searching the mutations. Methods The sequence of exon 10 and exon 13 in muta-tion hot spot region of FGFR3 gene in seven families was analyzed using polymerase chain reaction (PCR) and DNA sequenc-ing technology. Results The c.1138G>A missense mutation in exon 10 was found in 4 probands who were diagnosed as achon-droplasia (ACH), while this mutation was absent in their parents. The c.1620C>A missense mutation in exon 13 was found in one girl and her mother who both were diagnosed as hypochondroplasia (HCH) with mild symptoms. Neither mutation men-tioned above was found in the other two probands. Conclusions Through detecting the mutation in exon 10, exon 13 of FGFR3 gene, most patients of ACH or HCH can be finally diagnosed. However, it is necessary to perform the mutation screening on the other zones of FGFR3 gene and on other related genes for a few cases.

Thanatophoric dysplasia in a dichorionic twin confirmed by genetic analysis at the early second trimester: A case report and literature review

Thanatophoric dysplasia (TD) is caused by mutation of the gene that encodes fibroblast growth factor 3 (FGFR3). Owing to the poor prognosis for TD, prenatal diagnosis is critical to optimal perinatal management. We report here a case of TD in twin pregnancy, which was prenatally diagnosed by DNA analysis following amniocentesis at 15 weeks, and was managed by selective fetal termination. Prenatal ultrasonography and molecular analysis to detect TD-specific mutations enable accurate diagnosis of FGFR3-related TD in utero and appropriate obstetrical management at early gestation during twin pregnancy.

Clinical Experiences of Molecular Genetic Evaluation of Achondroplasia in Prenatal and Neonatal Cases

PURPOSE: The purpose of this study was to assess the characteristics of achondroplasia (ACH) diagnosed in fetuses or neonates and to evaluate the usefulness of a molecular genetic testing to confirm ACH. MATERIALS AND METHODS: The medical and ultrasonographic records of 16 pregnant women, who had molecular genetic testing for ACH performed on their fetus or neonate at the Cheil General Hospital between February 1999 and April 2013, were retrospectively analyzed. Detection of G1138A and G1138C mutations of the fibroblast growth factor receptor 3 (FGFR3) gene was accomplished by polymerase chain reaction - restriction fragment length polymorphism analysis. RESULTS: Of the eight fetuses and two neonates who were suspected of having ACH during pregnancy, four fetuses and one neonate was confirmed to have ACH and they all carried the heterozygous G1138A mutation. Out of 6 cases which had a history of ACH in prior pregnancies, three had genetic information for the previous fetuses while the other three did not. All six fetuses had no mutations at G380R. However, the one fetus of pregnant woman with non-confirmed ACH showed shortened long bone on ultrasound thereafter and the fetus was identified as having oto-spondylo-megaepiphyseal dysplasia after birth. CONCLUSION: Korean patients with achondroplasia have the heterozygous G1138A mutation that is most commonly defined in other countries. Molecular genetic evaluations of ACH are helpful not only for establishing diagnosis but for appropriate counseling with subsequent pregnancies.

Expression of Fibroblast Growth Factor Receptor 3 in the Recurrence of Non-Muscle-Invasive Urothelial Carcinoma of the Bladder

PURPOSE: The fibroblast growth factor receptor 3 (FGFR3) gene is known to be frequently mutated in noninvasive urothelial carcinomas of the bladder. In this study, we investigated the expression of FGFR3, Ki-67, and p53 in bladder cancers and the effects of expression on tumor recurrence. MATERIALS AND METHODS: Fifty-five cases of primary bladder cancer were examined by immunohistochemistry. The relationship of these markers with various clinicopathological factors, including recurrence, was assessed. RESULTS: Positivity for cytoplasmic FGFR3 (FGFR3-c) was associated with a lower cancer grade (p=0.022) and stage (p=0.011). Recurrence was more frequent in patients with a higher stage, negative FGFR3-c, and high Ki-67 expression. According to univariate analysis, predictors of recurrence-free survival included the following: age, stage, FGFR-c, Ki-67, and p53. However, none of these was independent from the other parameters in multivariate studies. CONCLUSIONS: The immunohistochemical expression of FGFR3 is not only one of the characteristic features of lower-grade and lower-stage urothelial carcinoma but also a possible marker in predicting disease recurrence.

The effect of pulsed electromagnetic fields on the proliferation of immunomagnetically separated human pre-cartilaginous stem cells / 中华物理医学与康复杂志

Objective To study the biological effects of pulsed electromagnetic fields (PEMFs) on the pro-liferation of immunomagnetically separated human precartilaginous stem cells (PSCs) in vitro. Methods The cells from an aborted fetus's metaphysis were digested using collagenase. The PSCs were isolated by magnetic cell sorting (MACS), then subcultured and amplified. Flow cytometry, immunohistochemistry, immunofluorescence and RT-PCR analysis were performed to identify the purified PSCs. The PSCs were stimulated by PEMFs at 50 Hz frequency and 1 mT intensity. Cell proliferation was measured at different time points using methyl thiazolyl tetrazolium ( MTT), and the cell growth curve was plotted. Flow cytometry was applied to measure the cell cycle and apoptosis. Results The PSCs were successfully cultured. There was fibroblast growth factor receptor-3 (FGFR-3) on their sur-face. Cell proliferation was promoted after 4 and 6 days of PEMF stimulation. The percentage of cells at the S phase was higher than in a control group. Early, late and total rates of apoptosis in the experimental group decreased signifi-cantly. Conclusion PEMFs can enhance the proliferation and inhibit the apoptosis of human PSCs, and it is possi-ble to cultivate the high density human PSCs in vitro.

Immunomagnetic Indirect Positive Sorting of Precartilaginous Stem Cells from Neonatal Rat / 华中科技大学学报（医学）（英德文版）

To investigate the technique of sorting high-purity precartilaginous stem cells from rat's perichondrium, neonatal rat's perichondrium cells suspensions were incubated with monoclone antibody of anti-fibroblast growth factor receptor-3 (anti-FGFR-3), and the labeled cells were separated from the suspension in the magnetic field by immuno-beads coated with the second antibody. Purityof the sorted neural stem cells was found to be 93.0 %-99.0 %, with living cells amounting to 80 %-85 %. The magnetic cell sorting system could effectively separate precartilaginous stem cells fromperichondrium cell suspensions.

The Effects of FGFR3 in Development Phase of Murine Small Intestine / 医学研究杂志

Objective To investigate the effect of fibroblast growth factor receptor 3(FGFR3) in development phase of murine small intestine.Methods Wild type mice and their littermate FGFR3+ mice were used to observe their morphology and proliferation characteristics of epithelia of small intestinine in development phase and changes of FGFR3 expression.Results Mutant mice had lower density and their villa were lower than those of the controls.But to the depth of crypt,the results were conversely.Cells in proliferation mainly located in intestine crypt.Mutant mice had more proliferation than the controls at every time point.Expression of FGFR3 was detected at birth(day 1),and the expression was maximal from day7 to day 21,decreasing rapidly thereafter to reach the relatively low at day35.And the expression of FGFR3 also located in intestine crypt,overlapping with that of Brdu.Conclusion FGFR3 improves formation of murine intestine crypt in development phase.

Detecting and Analyzing on Mutation of Fibroblost Growth Factor Recepter 3 Gene in A Congenital Achondroplasia Family / 实用儿科临床杂志

Objective To detect and analyze the mutation of fibroblast growth factor receptor 3(FGFR3) gene among a family with congenital achondroplasia(ACH).Methods Six blood samples of family member in this pedigree were cellected according to the informed consent process for genetic research,and 2 unralted healthy human blood sample were taken as controls.The mutation at nucleotide position 1 138 on FGFR3 gene was detected by using Polymerase chain reaction and single-strand conformation polymorphism(PCR-SSCP)and polyme-rase chain reaction and restriction endonuclease technology(PCR-RFLP) methods.Results Using PCR-SSCP method firstly,only the proband with ACH and his father in this family had the same abnormal band.The amplified products including 1 138 loci on FGFR3 gene further was analyzed by Sfe Ⅰ digestion,3 fragments including 164 bp,109 bp and 55 bp were detected in the proband and his father again,and the other members in the family and 2 controls just showed 164 bp band.It indicated that just 2 patients (proband and his father) showed heterozygous G→A transition mutation at nucleotide position 1 138 on the FGFR3 gene.The amplified products at 1 138 loci was also detected by MspⅠ digestion,just 1 band was observed in all members in this family and 2 controls.It showed that there was no G→C substitution at nucleotide position 1 138.Conclusions The G→A transition mutation at nucleotide position 1 138 in transmembrane domain of FGFR3 gene may be the main cause of achondroplasia in this family.In this pedigree,the proband showed's father a de novo mutation which was transferred to his child again.

Effect of FGFR3 transfection on CbfaⅠ and Collagen Ⅰ expression in mouse mesenchymal stem cells / 第三军医大学学报

Objective To explore the effect of fibroblast growth factor receptor 3(FGFR3)on the expressions of CbfaⅠ and Collagen Ⅰ in mesenchymal stem cells(MSCs)after Ad-FGFR3 transfection.Methods After Ad-FGFR3 and Ad-GFP were respectively transfected into mouse mesenchymal stem cells,the expression of FGFR3,CbfaⅠ and Collagen Ⅰ mRNAs and proteins were detected by RT-PCR and Western blot.Results The expressions of FGFR3,CbfaⅠ and Collagen Ⅰ were significantly higher in Ad-FGFR3 transfected MSCs than in Ad-GFP group.Conclusion FGFR3 can up-regulate the expression of CbfaⅠ and Collagen Ⅰ.

Genetical diagnosis in a congenital achondroplasia family / 中华内分泌代谢杂志

Objective To indentify the gene mutation of fibroblast growth factor receptor 3 (FGFR3) gene in a Chinese family with congenital achondroplasia (ACH). Methods The genomic DNA from 2 clinically diagnosed ACH patients and the other 4 members from the same family was prepared for PCR. The products of PCR were purified and then sequenced directly. Results Two patients with ACH in this family showed G-A transition mutation at nucleotide 1138 as heterozygotes. Conclusion The G-A transition mutation at nucleotide 1138 in transmembrane domain of FGFR3 gene seems to be the pathologic cause of this Chinese family with ACH.

Mutation at exon 10 of the fibroblast growth factor receptor 3 (FGFR3) in a fetus with thanatophoric dysplasia type I (TDI) / 대한산부인과학회잡지

Thanatophoric dysplasia (TD) is a sporadic lethal type of skeletal dysplasia featuring micromelia, decreased thoracic dimension and macrocephaly. To date, several kinds of mutation in fibroblast growth factor receptor 3 (FGFR3) has been identified in TD. We experienced a case of TD type I and underwent sequencing of the exon 7, 10 and the stop codon of FGFR3 to identify the type of mutation. TDI was diagnosed by the prenatal ultrasound at 25 weeks of gestation. The pregnancy was terminated and the diagnosis was confirmed by radiological and histologic examinations. The genomic DNA was extracted and the sequences of the exon 7, 10 and the stop codon of FGFR3 were amplified by PCR. The sequencing was performed for the each PCR products by dideoxyterminator method. The nucleotide transition from G to T was found in the nucleotide 1108, which is a part of the transmembrane domain, exon 10. To date, only one type of mutation (nucleotide 742) in the FGFR3 was identified in TD1 among Asian. This case firstly reveals the mutation of FGFR3 other than mutation at nucleotide 742 in TD1.