New recessive compound heterozygous variants of RP1L1 in RP1L1 maculopathy

AIM:To identify a maculopathy patient caused by new recessive compound heterozygous variants in RP1L1.METHODS:Comprehensive retinal morphological and functional examinations were evaluated for the patient with RP1L1 maculopathy.Targeted sequence capture array technique was used to screen potential pathologic variants.Polymerase chain reaction and Sanger sequencing were used to confirm the screening results.RESULTS:Fundus examination showed round macular lesions appeared in both eyes.Optical coherence tomography showed that the inner segment/outer segment continuity was disorganized and disruptive in the left eye,but it was uneven and slightly elevated in the right eye.Fundus autofluorescence showed patchy hyper-autofluorescence in the macula.Visual field examination indicates central defects in both eyes.Electroretinogram(ERG)and multifocal ERG showed no obvious abnormalities.Fundus fluorescein angiography in the macula showed obviously irregular hyper-fluorescence in the right eye and slightly hyper-fluorescence in the left eye.We found that the proband carried a missense variant(c.1972C>T)and a deletion variant(c.4717_4718del)of RP1L1,which were originated from the parents and formed compound heterozygous variants.Both variants are likely pathogenic according to the ACMG criteria.Multimodal imaging,ERG and detailed medical history are important diagnostic tools for differentiating between acquired and inherited retinal disorders.CONCLUSION:A maculopathy case with detailed retinal phenotype and new recessive compound heterozygous variants of RP1L1 is identified in a Chinese family,which expands the understanding of phenotype and genotype in RP1L1 maculopathy.

Complex heterozygous mutations in hereditary spherocytosis:A case report

BACKGROUND The aim of this study was to investigate the complex heterozygous mutations of ANK1 and SPTA1 in the same individual and improve our understanding of hereditary spherocytosis(HS)in children.We also hope to promote the application of gene detection technology in children with HS,with the goals of identifying more related gene mutations,supporting the acquisition of improved molecular genetic information to further reveal the pathogenesis of HS in children,and providing important guidance for the diagnosis,treatment,and prevention of HS in children.CASE SUMMARY A 1-year and 5-month-old patient presented jaundice during the neonatal period,mild anemia 8 months later,splenic enlargement at 1 year and 5 months,and brittle red blood cell permeability.Genetic testing was performed on the patient,their parents,and sister.Swiss Model software was used to predict the protein structure of complex heterozygous mutations in ANK1 and SPTA1.Genetic testing revealed that the patient harbored a new mutation in the ANK1 gene from the father and a mutation in the SPTA1 gene from the mother.Combined with the clinical symptoms of the children,it is suggested that the newly discovered complex heterozygous mutations of ANK1 and SPTA1 may be the cause,providing important guidance for revealing the pathogenesis,diagnosis,treatment,and promotion of gene detection technology in children with HS.CONCLUSION This case involves an unreported complex heterozygous mutation of ANK1 and SPTA1,which provides a reference for exploring HS.

Compound heterozygous mutations in tripeptidyl peptidase 1 cause rare autosomal recessive spinocerebellar ataxia type 7:A case report

BACKGROUND Spinocerebellar ataxia recessive type 7(SCAR7)is a rare clinical manifestation beginning in childhood or adolescence.SCAR7 is caused by tripeptidyl peptidase 1(TPP1)gene mutations,and presents with cerebellar ataxia,pyramidal signs,neurocognitive impairment,deep paresthesia,and cerebellar atrophy.CASE SUMMARY Here,we describe a 25-year-old female patient in China who presented with increasing difficulty walking,falling easily,shaking limbs,instability holding items,slurred speech,coughing when drinking,palpitations,and frequent hunger and overeating.Magnetic resonance imaging showed cerebellar atrophy.Whole exome sequencing detected two compound heterozygous mutations in the TPP1 gene:c.1468G>A p.Glu490Lys and c.1417G>A p.Gly473Arg.Considering the patient’s clinical presentation and genetic test results,we hypothesized that complex heterozygous mutations cause TPP1 enzyme deficiency,which may lead to SCAR7.CONCLUSION We report the first case of SCAR7 from China.We also identify novel compound heterozygous mutations in the TPP1 gene associated with SCAR7,expanding the range of known disease-causing mutations for SCAR7.

The Prevalence of Heterozygous Familial Hypercholesterolemia among Adult Filipino Patients with Dyslipidemia at Universidad de Santa Isabel Health Services Department: An Observational Descriptive Prospective Study

Objectives: It is to determine the prevalence of familial hypercholesterolemia (FH) among adult Filipino patients with dyslipidemia at Universidad de Santa Isabel Health Services Department in one year. Methods: An observational descriptive prospective study involves Filipino patients, aged 19 years and older, with dyslipidemia. The Dutch Lipid Network (DLN) Criteria was used to diagnose FH. Prevalence data and categorical variables were expressed as percentages, while continuous variables were reported as mean and standard deviations. Results: 529 patients were included in the study. 302 were females, and 227 were males. 180 (34%) scored Unlikely, 100 (19%) scored Probable, 185 (35%) scored Possible, and 64 (12%) were classified under Definite Familial Hypercholesterolemia. Most of the patients diagnosed with definite FH did not have diabetes, cerebrovascular disease (CVD), and coronary artery disease (CAD). The diagnosis was not affected by gender, BMI, smoking, and alcohol consumption. Hypertension was significantly correlated to the diagnosis of FH, as most of them were already hypertensive at diagnosis. It was noted that hypertension, diabetes, CVD, and CAD were seen at an earlier age among patients with definite FH. Conclusion: The prevalence of heterozygous FH at 12% among dyslipidemia patients and 1.3% among the general population was described for the first time in our region. This result should raise the awareness of our healthcare providers that FH, which is a major risk factor for premature CAD and CVD, exists, and early detection and management are important.

Mesenteric vein thrombosis in a patient heterozygous for factor Ⅴ Leiden and G20210A prothrombin genotypes

Mesenteric venous thrombosis(MVT)is a rare but life threatening form of bowel ischemia.It is implicated in 6%-9% of all cases of acute mesenteric ischemia.The proportion of patients with primary(or idiopathic)MVT varies from 0% to 49%,with a decrease in frequency secondary to more recent availability of newer investigations for hypercoagulability.The presence of factor Ⅴ Leiden(FVL)and prothrombin G20210A mutations(PGM)have been well documented in these cases.However,there have been scarce case reports describing MVT in heterozygotes of both these mutations occurring simultaneously and its implications on long term management.Our case describes acute MVT in a previously asymptomatic young patient with no prior history of venous thromboembolism.The patient was found to be heterozygous for FVL and PGM and treated with lifelong anticoagulation with warfarin(goal international normalized ratio:2-3)and avoidance of hormonal contraceptives.

Mice with a heterozygous Lrp6 deletion have impaired fracture healing

Bone fracture non-unions, the failure of a fracture to heal, occur in 10%-20% of fractures and are a costly and debilitating clinical problem. The Wnt/fl-catenin pathway is critical in bone development and fracture healing. Polymorphisms of linking low-density lipoprotein receptor-related protein 6 （LRP6）, a Wnt-binding receptor, have been associated with decreased bone mineral density and fragility fractures, although this remains controversial. Mice with a homozygous deletion of Lrp6 have severe skeletal abnormalities and are not viable, whereas mice with a heterozygous deletion have a combinatory effect with Lrp5 to decrease bone mineral density. As fracture healing closely models embryonic skeletal development, we investigated the process of fracture healing in mice heterozygous for Lrp6 （Lrp6~/-） and hypothesized that the heterozygous deletion of Lrp6 would impair fracture healing. Mid-diaphyseal femur fractures were induced in Lrp6~^- mice and wild-type controls （Lrp6~/~）. Fractures were analyzed using micro-computed tomography （~CT） scans, biomechanical testing, and histological analysis. Lrp6~/- mice had significantly decreased stiffness and strength at 28 days post fracture （PF） and significantly decreased BV/TV, total density, immature bone density, and mature area within the callus on day-14 and -21 PF; they had significantly increased empty callus area at days 14 and 21 PF. Our results demonstrate that the heterozygous deletion of Lrp6 impairs fracture healing, which suggests that Lrp6 has a role in fracture healing.

Dissection of QTLs for Yield Traits Using Near Isogenic Lines Derived from Residual Heterozygous Lines in Rice

Three residual heterozygous lines (RHLs) carrying heterozygous segments in the intervals RM587–RM225, RM204–RM6119 and RM6119–RM402 on the short arm of rice chromosome 6, respectively, were selected from a rice population derived from an RHL for the interval RM587–RM402. Ten maternal homozygotes, 10 paternal homozygotes and 20 heterozygotes were selected from each of the F2 populations derived from the three RHLs. The three sets of near isogenic lines (NILs) were grown to detect the grain yield per plant, number of panicles per plant, number of filled grains per panicle and 1000-grain weight. With analysis on the phenotypic differences among the three genotype groups in each NIL set and those among overlapping chromosome segment substitution lines, three QTLs for number of filled grains per panicle and two QTLs for grain yield per plant were resolved. They were located in the intervals ranging from 0.66 Mb to 2.49 Mb. The additive effect was higher than the dominance effect at each locus. The allele for increasing the trait value was derived from the paternal parent at qNFGP6-1, and from the maternal alleles at other QTLs. Based on the present study, an approach for constructing new genetic resource to facilitate fine mapping of QTLs in rice was proposed.

Novel compound heterozygous GPR56 gene mutation in a twin with lissencephaly:A case report

BACKGROUND Lissencephaly(LIS)is a malformation of cortical development with broad gyri,shallow sulci and thickened cortex characterized by developmental delays and seizures.Currently,20 genes have been implicated in LIS.However,GRP56-related LIS has never been reported.GRP56 is considered one of the causative genes for bilateral frontoparietal polymicrogyria.Here,we report a twin infant with LIS and review the relevant literature.The twins both carried the novel compound heterozygous GPR56 mutations.CASE SUMMARY A 5-mo-old female infant was hospitalized due to repeated convulsions for 1 d.The patient had a flat head deformity that manifested as developmental delays and a sudden onset of generalized tonic-clonic seizures at 5 mo without any causes.The electroencephalography was normal.Brain magnetic resonance imaging revealed a simple brain structure with widened and thickened gyri and shallow sulci.The white matter of the brain was significantly reduced.Patchy long T1 and T2 signals could be seen around the ventricles,which were expanded,and the extracerebral space was widened.Genetic testing confirmed that the patient carried the GPR56 gene compound heterozygous mutations c.228delC(p.F76fs)and c.1820_1821delAT(p.H607fs).The unaffected father carried a heterozygous c.1820_1821delAT mutation,and the unaffected mother carried a heterozygous c.228delC mutation.The twin sister carried the same mutations as the proband.The patient was diagnosed with LIS.CONCLUSION This is the first case report of LIS that is likely caused by mutations of the GPR56 gene.

Heterozygous carriers of galactocerebrosidase mutations that cause Krabbe disease have impaired microglial function and defective repair of myelin damage

This review addresses two puzzling findings related to mutations in galactocerebrosidase （GALC） that cause Krabbe disease （KD）, a severe lysosomal storage disorder characterized by extensive myelin damage in children with mutations in both GALC alleles. First, heterozygous carriers of KD-causing mutations, which include the biological parents of children with KD, exhibit increased risk for developing other diseases. Second, variants in the GALC locus increase the risk of developing multiple sclerosis （MS）, another disease characterized by extensive myelin damage. What explains these correlations？ In studies on cuprizone-induced myelin damage in heterozygous （GALC＋/–） mice carrying one copy of a mutation that causes KD-like disease, the extent of damage was similar in GALC＋/– and wild-type （WT） mice. In contrast, GALC＋/- mice had striking defects in repair of cuprizone-induced damage. We further found unexpected microglial defects in myelin debris clearance and in the ability to up-regulate the Trem2 microglial protein critical for debris uptake. These defects were rescued by exposure to a lysosomal re-acidifying drug discovered in our studies on KD, and which provides multiple clinically relevant benefits in the twitcher （GALC＋/–） mouse model of KD. Thus, heterozygous GALC mutations cause effects on biological function that may help to understand the increased disease risk in heterozygous carriers of such mutations and to understand why GALC variations increase the risk of MS. Our findings indicate that while some genetic risk factors may contribute to complex diseases by increasing the risk of tissue damage, others may do so by compromising tissue repair.

Characteristics and Clinical Implication of UGT1A1 Heterozygous Mutation in Tumor

Background:The literature recommends that reduced dosage of CPT-11 should be applied in patients with UGT1 A1 homozygous mutations,but the impact of UGT1 A1 heterozygous mutations on the adverse reactions of CPT-11 is still not fully clear.Methods:A total of 107 patients with UGT1 A1 heterozygous mutation or wild-type,who were treated with CPT-11 from January 2018 to September 2021 in Peking University Third Hospital,were retrospectively enrolled.The adverse reaction spectra of patients with UGT1 A1*6 and UGT1 A1*28 mutations were analyzed.Adverse reactions were evaluated according to National Cancer Institute Common Terminology Criteria for Adverse Events(NCI-CTCAE) 5.0.The efficacy was evaluated according to Response Evaluation Criteria in Solid Tumors(RECIST) 1.1.The genotypes of UGT1 A1*6 and UGT1 A1*28 were detected by digital fluorescence molecular hybridization.Results:There were 43 patients with UGT1 A1*6 heterozygous mutation,26 patients with UGT1 A1*28 heterozygous mutation,8 patients with UGT1 A1*6 and UGT1 A1*28 double heterozygous mutations,61 patients with heterozygous mutation at any gene locus of UGT1 A1*6 and UGT1 A1*28.Logistic regression analysis showed that the presence or absence of vomiting(P=0.013) and mucositis(P=0.005) was significantly correlated with heterozygous mutation of UGT1 A1*28,and the severity of vomiting(P<0.001) and neutropenia(P=0.021) were significantly correlated with heterozygous mutation of UGT1 A1*6.In colorectal cancer,UGT1 A1*6 was significantly correlated to diarrhea(P=0.005),and the other adverse reactions spectrum was similar to that of the whole patient cohort,and efficacy and prognosis were similar between patients with different genotypes and patients treated with reduced CPT-11 dosage or not.Conclusion:In clinical use,heterozygous mutations of UGT1 A1*6 and UGT1 A1*28 are related to the risk and severity of vomiting,diarrhea,neutropenia and mucositis in patients with Pan-tumor and colorectal cancer post CPT-11 therpy.In colorectal cancer,UGT1 A1*6 is significantly related to diarrhea post CPT-11 use,efficacy and prognosis is not affected by various genotypes or CPT-11 dosage reduction.

Child with adenylosuccinate lyase deficiency caused by a novel complex heterozygous mutation in the ADSL gene:A case report

BACKGROUND Adenylosuccinate lyase(ADSL)deficiency is a rare autosomal-recessive defect of purine metabolism caused by mutation of the ADSL gene.It can cause severe neurological impairment and diverse clinical manifestations,including epilepsy.CASE SUMMARY Here,we describe a 3-year-old Chinese boy who had both psychomotor retardation and refractory epilepsy.Magnetic resonance imaging showed myelin hypoplasia.Electroencephalography findings supported a diagnosis of epilepsy.Whole-exon sequencing revealed the presence of a novel complex heterozygous mutation in the ADSL gene:The splicing mutation c.154-3C>G and the missense mutation c.71C>T(p.Pro24Leu).Considering the patient’s clinical presentation and genetic test results,the complex heterozygous mutation was predicted to prevent both ADSL alleles from producing normal ADSL,which may have led to ADSL deficiency.Finally,the child was diagnosed with ADSL deficiency.CONCLUSION We identified a novel complex heterozygous mutation in the ADSL gene associated with ADSL deficiency,thus expanding the known spectrum of pathogenic mutations that cause ADSL deficiency.Additionally,we describe epilepsy that occurs in patients with ADSL deficiency.

Digenic heterozygous mutations in EYS/LRP5 in a Chinese family with retinitis pigmentosa

Dear Editor,I am Dr.Ji-Hong Wu,from the Department of Ophthalmology,Eye＆ENT Hospital of Fudan University,China.I write to present a case report of retinitis pigmentosa（RP）caused by novel digenic heterozygous mutations in a Chinese family.

Effect of LDL-apheresis on plasma lipids, chitotriosidase and anti-oxLDL antibodies in heterozygous familial hypercholes-terolemia

Forty four consecutive subjects aged 29-58 years (21 males and 23 females) with a clinical diagnosis of heterozygous familial hypercholesterolemia periodically treated every 30 days with LDL-apheresis for statin resistance, were enrolled in this study. A lipid profile was obtained immediately before starting LDL-apheresis, a second profile was obtained within four hours after LDL-apheresis. Chit activity and anti-oxLDL levels were determined with appropriate methods in all patients before and after LDL- apheresis. Total cholesterol, LDL-cholesterol, HDL- cholesterol and triglycerides decreased significantly after LDL-apheresis, while the variations of Chit activity and anti-oxLDL were not significant after LDL-apheresis. The correlation between Chit and total cholesterol was negative (r= –0.44 and –0.50 res- pectively) before and after LDL-apheresis as between Chit and LDL-cholesterol (r= –0.45 and –0.55 respectively). Anti-oxLDL concentration before and after LDL-apheresis positively correlated with Chit activity (r= 0.52 and r = 0.63 respectively), negatively with total cholesterol (r= –0.33 and r = –0.35 res- pectively) and with LDL (r = –0.32 and r = –0.21 respectively). We think that removing LDL with LDL-apheresis the anti-oxLDL/oxLDL ratio could increase and the excess of anti-oxLDL could induce macrophage activation through the surface Fc receptors. Alternatively with high levels of LDL- cholesterol, the deposition of foam cells represent the characteristic evolution of atherosclerosis process. Macrophage activation in the heterozygous familial hypercholesterolemia could represent an attempt for re-modeling the vessel wall, reducing the growth of lipid plaques.

Delivery Outcome in Women with Major Sickle Cell Syndrome: A Comparative Study of the Homozygous Forms “SS” versus the Heterozygous “SC”

Objectives: To determine the prevalence of women who delivered in the two major sickle cell syndromes, “SS” and “SC”, and to identify maternal and early neonatal prognosis inherent to each form. Material and Methods: This is a comparative, descriptive and retrospective cross-sectional study of 226 files of women carrying major sickle cell syndrome (66 cases of “SS” form versus 160 cases of “SC” form), collected from May 2008 to May 2013 at the Gynecology and Obstetrics Clinic of the Sylvanus Olympio’s University Hospital of Lomé. Data were processed by Epi Info 6 software. For comparison of variables, the Chi-2 test of Fisher with significance as p < 0.05 has been used, so is the calculation of Odds Ratio with its confidence interval at 95%. Results: Carriers of the two major sickle cell syndromes represent 0.8% of all the deliveries during the study period. Caesarean section, especially prophylactic one, was the dominant mode of delivery. The SS forms have been exposed to have more vaso-occlusive crises (22.7% vs. 13.1%;p = 0.04, OR = 0.31), more blood transfusion (57.6% vs. 29.4%;p = 7 × 10-5, OR = 3.2) and more puerperal infections (p < 0.05). Acute chest syndrome was not related to any of the two forms of sickle cell disease (13.6% vs. 8.1%;p = 0.15). The maternal mortality rate and the perinatal mortality among SS form against SC form were not significant (respectively 15.1% vs. 8.7%;20.9% vs. 17.1%). Conclusion: Maternal and fetal complications were present in both forms of major sickle cell syndrome but the "SS" form gave exposure to greater maternal morbidity. Resuscitative measures in adults and newborns should be reinforced at the delivery time of these “at-risk-pregnancies”.

"Heterozygous Treatment"Method to Improve the Biased Status of the Damp-Heat Constitution and Symptom Integral:Randomized Controlled Trial

Objective:To observe the clinical effect of"heterozygous treatment"intervening the damp-heat constitution.Method:One hundred and six cases with damp-heat constitution were randomly divided into the observation group and control group,fifty-three cases for each group.Lianpu drink was given to the two groups,and the observation group was treated with scraping,acupuncture,cupping,constitution care and popularization of constitution science for"heterozygous treatment"based on the control group.70 days later,"constitution classification and determination table of traditional Chinese medicine"was used to determine,and statistics was applied to analyze the change of the symptoms of the two groups before and after the intervention.Results:in the observation group,compared to before the intervention,symptoms like dirty and oily complexion,yellow greasy tongue,bitter taste,dullness and scanty dark urine were significantly improved(P<0.05),and improvement of the above symptoms was greater than the control group(P<0.05).Conclusion:There is no significant difference between the two groups(P>0.05).Conclusion:the"heterozygous treatment"method can significantly improve the clinical symptoms of people with damp-heat constitution,with a better role in regulating.

Prevalence of Cardiovascular Risk Factors and Heterozygous Familial Hypercholesterolemia in Premature Atherosclerotic Patients Presenting with Acute Coronary Syndrome

Background: Heterozygous familial hypercholesterolemia is an autosomal dominant genetic disorder with an estimated prevalence of 1/200 - 1/500 in the general population. Early identification of patient with familial hypercholesterolemia is important, because appropriate treatment may reduce the risk of premature atherosclerosis. Objective: Assessment of the prevalence of different modifiable cardiovascular risk factors and clinical diagnosis of heterozygous familial hypercholesterolemia. Methods: One hundred patients were enrolled, included young patients (males less than 50 years and females less than 60 years old) presented with first attack of acute coronary syndrome either ST elevation myocardial infarction (STEMI), non ST elevation myocardial infarction (NSTEMI) or unstable angina (UA). All patients were subjected to full history taking, general and local examination, Electrocardiogram, transthoracic Echocardiography, laboratory investigations, coronary angiography and Dutch score calculation for familial hyperlipidemias. Results: The mean level of serum cholesterol among studied group was 268.31 ± 59.33, HDL-C was 39.63 ± 7.52, LDL was 192.27 ± 60.61 and TG was 180.10 ± 39.64. With application of Dutch score, 20% of patients diagnosed definite familial hypercholesterolemia with Dutch score > 8. Twenty-six percent of patients diagnosed as probable familial hypercholesterolemia with Dutch score 6 - 8. Thirty-nine percent patients diagnosed as possible familial hypercholesterolemia with Dutch score 3 - 5 and 15% of patients were unlikely familial hypercholesterolemia with Dutch score < 3 with significant correlation between Dutch score and age, total cholesterol, LDL-C, serum creatinine. Conclusion: Familial hypercholesterolemia (FH) is one of the most common serious genetic disorders of cholesterol metabolism. The early identification of heterogynous FH patients is crucial to start an effective prevention strategy.

Genetic Analysis of Two Novel GPI Variants Disrupting H Bonds and Localization Characteristics of 55 Gene Variants Associated with Glucose-6-phosphate Isomerase Deficiency

Objective:Glucose-6-phosphate isomerase(GPI)deficiency is a rare hereditary nonspherocytic hemolytic anemia caused by GPI gene variants.This disorder exhibits wide heterogeneity in its clinical manifestations and molecular characteristics,often posing challenges for precise diagnoses using conventional methods.To this end,this study aimed to identify the novel variants responsible for GPI deficiency in a Chinese family.Methods:The clinical manifestations of the patient were summarized and analyzed for GPI deficiency phenotype diagnosis.Novel compound heterozygous variants of the GPI gene,c.174C>A(p.Asn58Lys)and c.1538G>T(p.Trp513Leu),were identified using whole-exome and Sanger sequencing.The AlphaFold program and Chimera software were used to analyze the effects of compound heterozygous variants on GPI structure.Results:By characterizing 53 GPI missense/nonsense variants from previous literature and two novel missense variants identified in this study,we found that most variants were located in exons 3,4,12,and 18,with a few localized in exons 8,9,and 14.This study identified novel compound heterozygous variants associated with GPI deficiency.These pathogenic variants disrupt hydrogen bonds formed by highly conserved GPI amino acids.Conclusion:Early family-based sequencing analyses,especially for patients with congenital anemia,can help increase diagnostic accuracy for GPI deficiency,improve child healthcare,and enable genetic counseling.

Compound heterozygous mutation in two unrelated cases of Chinese spinal muscular atrophy patients

Background Infantile proximal spinal muscular atrophy （SMA） is a common autosomal recessive neuromuscular disorder. Approximately 90-95% cases of SMA result from homozygous deletion of survival motor neuron gene 1（SMN1） and 5% cases are caused by compound heterozygous mutation （a SMN1 deletion on one allele and a subtle mutation on the other allele）.Methods In this research, two unrelated patients were clinically diagnosed according to the criteria of proximal SMA. Genetic diagnosis was performed to detect the homozygous deletion of exon 7 of SMN1 by PCR-restriction fragment length polymorphism （RFLP） and genomic sequencing. Multiplex ligation-dependent probe amplification （MLPA） analysis was carried out to measure copy numbers of SMN1, SMN2 and neuronal apoptosis inhibitor protein （NAIP） in the patients. Further sequencing of SMN1allele-specific PCR （AS-PCR） and SMN1 clones were also performed to analyze the point mutation of SMN1 gene. Additionally,the pedigree analysis of these two families was carried out to identify the transmission of the mutation.Results The inconsistent results using PCR-RFLP and genomic sequencing showed homozygous deletion of exon 7 of SMN1 and heterozygous deletion accompanied with a suspicious mutation in SMN1 gene, respectively. MLPA analysis of these two cases exhibited one SMN1 copy deletion. One identical c.863G〉T （p. Arg288Met） mutation was found in two cases by sequencing the SMN1 clones, which confirmed that both cases were SMA compound heterozygotes. One case showed partial conversion to form hybrid SMN （SMN2 17/SMN1 E8） identified by clones sequencing and another case carrying 3 SMN2 implied complete conversion from SMN1 to SMN2.Conclusion p. Arg288Met is more a disease-causing mutation than a polymorphism variation, and children with this mutation may have more severe phenotypes.

R158Q and G212S,novel pathogenic compound heterozygous variants in SLC12A3 of Gitelman syndrome

The dysfunction of Na^(+)-Cl^(−)cotransporter(NCC)caused by mutations in solute carrier family12,member 3 gene(SLC12A3)primarily causes Gitelman syndrome(GS).In identifying the pathogenicity of R158Q and G212S variants of SLC12A3,we evaluated the pathogenicity by bioinformatic,expression,and localization analysis of two variants from a patient in our cohort.The prediction of mutant protein showed that p.R158Q and p.G212S could alter protein’s three-dimensional structure.Western blot showed a decrease of mutant Ncc.Immunofluorescence of the two mutations revealed a diffuse positive staining below the plasma membrane.Meanwhile,we conducted a compound heterozygous model—Ncc^(R156Q/G210S)mice corresponding to human NCC R158Q/G212S.Ncc^(R156Q/G210S)mice clearly exhibited typical GS features,including hypokalemia,hypomagnesemia,and increased fractional excretion of K^(+)and Mg^(2+)with a normal blood pressure level,which made Ncc^(R156Q/G210S)mice an optimal mouse model for further study of GS.A dramatic decrease and abnormal localization of the mutant Ncc in distal convoluted tubules contributed to the phenotype.The hydrochlorothiazide test showed a loss of function of mutant Ncc in Ncc^(R156Q/G210S)mice.These findings indicated that R158Q and G212S variants of SLC12A3 were pathogenic variants of GS.

Compound heterozygous variants in POR gene identified by whole-exome sequencing in a Chinese pedigree with cytochrome P450 oxidoreductase deficiency

Importance:Cytochrome P450 oxidoreductase deficiency (PORD) is a rare disease exhibiting a variety of clinical manifestations.This condition specifically leads to disordered steroidogenesis,which can affect the development of the reproductive system,skeleton,and other parts of the body.The severe form of PORD is difficult to differentiate with Antley-Bixler syndrome (ABS).The genetic characters and clinical evaluation of PORD are still unclear in China.Objective:To perform an exome analysis and identify the pathogenic cause in order to assist clinicians to obtain a proper evaluation on the genetic condition.Methods:The proband underwent detailed physical evaluations.DNA of the proband and his parents was isolated and whole-exome sequencing (WES) was performed.Variants were analyzed and evaluation according to the ACMG guideline.Results:A 1-year-old Chinese boy with midface hypoplasia,choanal stenosis,multiple joint contractures,micropenis and right cryptorchidism was misdiagnosed with Crouzon syndrome.By trio-whole-exome sequencing,we identified an unreported compound heterozygous mutation (c.667C＞T,p.R223* and c.1370G＞A,p.R457H) in POR in the proband.This mutation was inherited from healthy heterozygous parents,supporting the diagnosis of PORD,which was further confirmed by biochemical characteristics.Interpretation:We have identified a pathogenic variant with an unreported compound heterozygous POR mutation,which expands the clinical and genetic spectra of PORD and emphasizes the usefulness of WES for genetic diagnosis.