Knee osteoarthritis (OA) is one of the most prevalent disorders in elderly population. Among various therapeutic alternatives, we employed stromal vascular fraction (SVF), a heterogeneous cell population, to regenerate damaged knee cartilage. OA patients were classified on the basis of age, gender, body mass index (BMI), and x-ray-derived Kellgren–Lawrence (KL) grade. They were treated with SVF and followed-up for 24 months. Visual analogue scale (VAS) and Western Ontario and McMaster Universities Osteoarthritis (WOMAC) Index were used to determine treatment efficacy. Cartilage healing was assessed using the MRI-based Outerbridge score (OS) and evaluation of bone marrow edema (BME) lesions, while a placebo group was used as a control. Time- and KL-dependent changes were also monitored. We observed a decreasing trend in VAS score and WOMAC index in the SVF-treated group up to 24 months, as compared with the placebo group. Besides, a significant increase and decrease in Lysholm and OS, respectively, were observed in the treatment group. Compared with the values before treatment, the greatly reduced WOMAC scores of KL3 than KL2 groups at 24 months, indicate more improvement in the KL3 group. Highly decreased BME in the treated group was also noted. In conclusion, the SVF therapy is effective in the recovery of OA patients of KL3 grade in 24 months. Knee osteoarthritis (OA) is one of the most common progressive joint disorders, especially among elderly population in the United States and other developed countries ,2,3. CarAn alternative surgical technique, the autologous chondrocyte implantation (ACI), has been recently used to overcome the limitations associated with the previously mentioned surgical techniques. ACI is a common surgical intervention to promote healing of cartilage injuries in OA ,29. HoweStromal vascular fraction (SVF) is a heterogeneous population of various immune, precursor, progenitor, and stem cells. SVF is considered to be as equal as or sometimes more effective than ADSCs; therefore, it provides other functional advantages, such as structural support, over ADSCs ,34,35,36This study was an open-label, single-center, non-randomized, placebo-controlled, phase I/II clinical trial to evaluate the improvement in knee pain and knee function, as well as cartilage restoration. The 33 patients enrolled in the study were deliberately allocated to two groups, which were designated arthroscopic microfracture treatment only and arthroscopic microfracture treatment combined with SVF injection. Observation and follow-up data were recorded after 12 and 24 months. The eligibility criteria included: osteoarthritic knee joint with KL grades 2–3 and age >38 years. Patients meeting the following criteria were excluded: autoimmune or inflammatory disease, infection requiring parenteral administration of antibiotics, serious internal disorders, corticosteroids or viscosupplements injection into the affected knee within the past 3 months, and stiffness due to previous severe injury. The protocol was approved by the Viet Nam Ministry of Health (No. 2288/QDBYT) and the Ethical Committee in Biomedical Research of Van Hanh General Hospital (No. 90-084/QD-BVVH). Patients participating in this research provided an informed consent, in accordance with the Declaration of Helsinki.Lipoaspirates were harvested from patients’ lower abdomen by a standard liposuction technique. Briefly, through incision, a solution of tumescent lidocain, 250 mL of normal saline, 0.9% and 0.2 mL of 1:1000 epinephrine was injected in the subcutaneous fat. Thereafter, 50–100 mL of lipoaspirate was collected through Triport Harvester Cannula , and a 60 mL Luer-lock syringe. The SVF from the lipoaspirate was isolated by means of collagenase digestion  and the ADSC Extraction Kit  approved by the Viet Nam Ministry of Health. The SVF was then washed thrice with sterile PBS to remove collagenase. Finally, the SVF was diluted with normal saline 0.9% to obtain 6 mL of solution containing 90–120 million cells to administer in each knee joint.Spinal anesthesia for knee arthroscopy was done by using 2 mL (5 mg/mL) bupivacaine hydrochloride. The debris, crystal, and synovitis were removed, and microfracture holes were placed 3–4 mm apart by the arthroscopy microfracture technique, as described by Steadman et al . After aPatients were monitored in the hospital for one week post-arthroscopy. After this, patients were followed for 24 months. Clinical manifestations such as pain, stiffness, and functional mobility were substantially recorded. Western Ontario and McMaster Universities Arthritis Index (WOMAC) , Lysholmt-test, using SPSS-22 , and p values <0.05 were considered statistically significant.The data are expressed as the mean ± SD. The comparisons between groups were performed by one-way analysis of variance (ANOVA) and The study was conducted from September 2014 to June 2017 at Van Hanh Hospital, Ho Chi Minh city, Vietnam. The overall schematic is illustrated in Eighteen patients who satisfied the exclusive and inclusive criteria were selected to receive the treatment of SVF, a heterogeneous cell population containing mesenchymal progenitor/stem cells, preadipocytes, endothelial cells, pericytes, T cells, and M2 macrophages . The demThe patients were classified on the basis of their age, gender, BMI, and KL grade . In genep < 0.05). Further, as compared to the placebo group, a sharp decreasing trend in the VAS score of the treatment group was observed up to 24 months. The VAS score in the treated group continuously reduced after 12 and 24 months. Specifically, compared to the mean VAS score at 12 months, the score at 24 months was significantly reduced . On the contrary, the score of the placebo group after 12 and 24 months increased from 4.9 ± 2 to 5.9 ± 2.47, but this difference was not significant. A similar trend was also observed for the WOMAC score in the placebo group, which was significantly decreased after 12 months of treatment . However, a significant increase was observed thereafter at 24 months . Meanwhile, the WOMAC score in the treated group decreased sharply after 12 months  and further declined significantly to 11.1 ± 11.9 at 24 months . Overall, at 24 months, both VAS and WOMAC scores in the placebo and treatment groups diminished compared with the scores before treatment. However, the decreasing trend in the treatment group was larger than in the placebo group, which is indicative of improvement after SVF therapy.VAS is a reliable scale for the assessment of pain in osteoarthritic condition , whereasp < 0.05) (p < 0.05), implying an improvement in knee function.The Lysholm Knee Scale is another recommended measure of knee function . As per  < 0.05) C. The reMRI results showed that after 24 months of treatment, bone marrow edema was decreased in both the placebo and the SVF treatment groups; however, the decrease in bone marrow edema in the SVF treatment group was larger (22 mm vs. 8 mm) than in the placebo group (20 mm vs. 12 mm) A. Similap < 0.05). Taken together, the OS score of the treated group clearly decreased, while that of the placebo group displayed nearly no change.The level of cartilage injury was measured by the Outerbridge score (OS) . The OS p < 0.05). Interestingly, compared to the placebo, the BME length before SVF treatment (2.4 ± 0.34 mm) was significantly larger than after 12 and 24 months of treatment . On the whole, these results indicate a reduction in the formation of BME-like lesions after SVF treatment. BME-like lesions are also associated with the pathogenesis of osteoarthritis and are characterized by histologic abnormalities such as bone marrow necrosis and fibrosis, in addition to trabecular abnormalities . TherefoThe X-ray image-derived KL grading scale is a gold standard for determining the severity of OA, on the basis of which, the total OA patients were divided into KL2 and KL3 groups . FurtherSimilarly, after treatment of KL2- and KL3-grade patients, differences in the WOMAC scores between the two groups were observed . The WOMThe impact of KL-OA grades on the Lysholm score is represented in The comparative profile of cartilage injury, as measured by the OS score in KL2 and KL3 patients after treatment, is represented in SVF contains a heterogeneous cell population of progenitor cells and ADSCs, which possess enhanced therapeutic potential against immune disorders, degenerative tissue pathologies, and other ischaemic conditions . The comp < 0.05). These data establish the role of SVF in improving the BME score which is used as an indicator of knee OA progression and is characterized by increased accumulation of fluid [Further, the level of cartilage injury was assessed on the MRI-based OS score. An increase in OS score represents a loss of cartilage thickness. In this study, initially there was no significant difference between the OS scores of the treatment and placebo groups; however, a significant decrease in OS score was observed in the treatment group compared to the placebo group after 24 months of treatment , endothelial cells (ECs), macrophages, smooth muscle cells, lymphocytes, pericytes, and pre-adipocytes ,66. The Besides these therapeutic activities, the regenerative ability of SVF may be due to ADSCs differentiation potential into chondrocytic and osteocytic cells lineages. EPCs may also induce angiogenesis by releasing growth factors such as vascular endothelial growth factor (VEGF) and insulin-like growth factor-1 (IGF-1) . MacrophOn the basis of the improvements observed in treated patients during follow-up and the behavior of the placebo group, our study revealed a trend toward a better efficacy of SVF with the microfracture method for OA treatment over a period of two years. We also inferred that the SVF therapy is more effective in KL 3-grade OA patients compared with KL 2-grade OA patients.


B3GLCT gene, of which c.660+1G>A (rs80338851) was previously associated with the phenotype of Peters plus syndrome (PPS), while the second, NM_194318.3:c.755delC (p.T252fs), in exon 9 of the same gene was noted for the first time. PPS, a very rare subtype of ASD, is a glycosylation disorder, where the dysfunctional B3GLCT gene product, O-fucose-specific β-1,3-glucosyltransferase, is ineffective in providing a noncanonical quality control system for proper protein folding in cells. Our study expands the mutation spectrum of the B3GLCT gene related to PPS. We suggest that the implementation of automatic text mining tools in combination with careful variant filtering could help translate sequencing results into diagnosis, thus, considerably accelerating the diagnostic process and, thereby, improving patient management.Anterior segment dysgenesis (ASD) encompasses a spectrum of ocular disorders affecting the structures of the anterior eye chamber. Mutations in several genes, involved in eye development, are implicated in this disorder. ASD is often accompanied by diverse multisystemic symptoms and another genetic cause, such as variants in genes encoding collagen type IV. Thus, a wide spectrum of phenotypes and underlying genetic diversity make fast and proper diagnosis challenging. Here, we used AMELIE, an automatic text mining tool that enriches data with the most up-to-date information from literature, and wANNOVAR, which is based on well-documented databases and incorporates variant filtering strategy to identify genetic variants responsible for severely-manifested ASD in a newborn child. This strategy, applied to trio sequencing data in compliance with ACMG 2015 guidelines, helped us find two compound heterozygous variants of the  In recent years, next-generation sequencing technology (NGS) and access to databases containing sequencing data from patients with eye disorders have accelerated research in ophthalmic molecular genetics. However, still the most difficult and time-consuming part of the work is data analysis and clinical interpretation of reported variants, especially in cases with blended phenotype, when the decision on major clinical feature, crucial for diagnostic process is difficult to make. Additionally, due to massive sequencing data generation and relatively restricted curation, available databases, within reliable information contain false variant pathogenicity attributions. Thus, rigorous variant interpretation, as recommended by the American College of Medical Genetics and Genomics (ACMG) , followeCYP1B1, BMP4, FOXC1, PAX6, FOXE3, NDP, SLC4A11, HCCS, PITX2, PITX3, LMX1B, and PXDN [COL4A1 and COL4A2 [Anterior segment dysgenesis (ASD) encompasses a spectrum of ocular disorders affecting the structures of the anterior eye chamber, including the iris, lens, cornea, and the anterior chamber angle. Clinical manifestations include corneal opacities, cataracts, and iridocorneal adhesions. ASD poses a high risk of the patient developing early onset and aggressive glaucoma resulting from dysregulation of aqueous humor flow, high intraocular pressure (IOP), and death of retinal ganglion cells . Disordeand PXDN ,6,7,8, sd COL4A2 ,10, ofteDue to the heterogeneous clinical manifestation of the syndrome, reflecting the complex genetic background, classification, and proper diagnosis of ASD is still challenging. Here, we report a strategy for NGS data analysis in order to find the genetic cause of a rare combination of symptoms in a pediatric patient with a set of ocular disorders, cleft lip and high-arched palate, using data from DNA sequencing of coding regions of 4813 genes in both parents and the affected child.We demonstrate a variant filtering strategy combined with information on the observed phenotype coded in Human Phenotype Ontology (HPO) terms and information enrichment by literature mining with the AMELIE   tool andMedian) and PAPP-A, 0.634 MoM. The first trimester combined with prenatal screening yielded a risk of 1:149 for trisomy 21, 1:7780 for trisomy 18, and 1:7447 for trisomy 13.The 30-year-old pregnant patient was referred for the first-trimester testing due to three previous miscarriages and advanced maternal age. The parents had one healthy child. Screening involved testing for trisomies 21, 18, and 13, ultrasonography, measurement of maternal serum free β-human chorionic gonadotropin (β-hCG), and pregnancy-associated plasma protein-A (PAPP-A) levels, according to the Fetal Medicine Foundation recommendations. Both biochemical parameters were measured on the DELFIA Xpress analyzer (Perkin Elmer). The mother denied medication history or infection during pregnancy. Moreover, no familial predisposition to any disease was identified. Routine fetal scanning at 13 weeks of gestation showed a cleft lip and palate and increased nuchal translucency . Nuchal Having detected a fetal defect and correlated it with an indirect risk group, an invasive diagnostic procedure was suggested to determine the fetal karyotype. Amniotic fluid was aspirated during amniocentesis on week 15, day 4 of pregnancy. Following amniocyte culture, a karyotype of a normal male  was obtained at a resolution of 550 bands. The couple was then referred for genetic counseling for further etiological investigation.The second ultrasonographic examination performed at the 20th week of gestation, as per the recommendations of the Polish Society of Gynecologists and Obstetricians, confirmed the presence of bilateral cleft lip and palate.The patient (mother) was hospitalized with intrauterine growth restriction in the 28th week of gestation. In view of abnormal flow in the umbilical artery and the middle cerebral artery, as well as deteriorating fetal well-being, it was decided to terminate the pregnancy and perform a cesarean section.Physical examination of the fetus revealed bilateral cleft alveolar processes, cleft lip, and high-arched palate. Rhizomelic proximal extremities and brachydactyly were noted. Microphthalmia and bilateral corneal opacities were observed during ophthalmic examination.Visual acuity in both eyes was determined as dubious light perception. Examination under general anesthesia resulted in the following findings .Right eye (RE): Cornea dimensions 8 x 8.5 mm; IOP 10 mmHg; central corneal leucoma, subsequent to ulceration with ingrowth of blood vessels. Translucent peripheral cornea revealed flattened anterior eye chamber, medium-width iris with persistent pupillary membrane, and lens adhesion to the posterior corneal surface. A visible fragment of the lens was translucent with pink reflection from the eye fundus. Further details were hard to evaluate.Left eye (LE): Cornea dimensions 8 × 8.5 mm; IOP 12 mmHg, partial central leucoma. The peripheral part of the cornea was translucent with a persistent pupillary membrane. The pupillary dilatory response was normal and some peripheral blood vessels of the retina could be visualized.The anatomical axial length of the eyeballs in ultrasonographic (USG) evaluation was 18 mm (RE) and 17.5 mm (LE) in projection AB . Diagnostic USG of the posterior part of the eyes revealed no pathology in RE and a small floater above an optic nerve disk in LE.The patient remained under ophthalmic supervision. At the age of four months, central thinning of the corneal leucoma of the RE, threatening perforation and secondary glaucoma were determined in the RE. Corneal thickness in the RE was 826 µm in the periphery but only 158–380 µm in the center while being 855 µm in the entire LE. IOP was 37 mmHg in the right and 17 mmHg in the LE. Amniotic membrane transplantation to protect the cornea and cyclocryotherapy to lower IOP were performed in the RE. Topical IOP lowering medications were started and the patient was followed for local status and IOP. In spite of ongoing pharmacological treatment, another round of amniotic membrane transplantation and cyclocryotherapy had to be performed at the age of six months. Stabilization of the IOP was achieved at the level of 26 mmHg in the right and 18 mmHg in the LE. The patient remains under ophthalmic supervision with permanent pharmacological medications to prevent glaucoma of the RE.On average, 9.8 million reads were sequenced per sample, and approximately 8.9 million reads (91.1%) were mapped on the reference using our custom pipeline. A high depth of coverage (at least 20×) was maintained for 86.9% of the targeted region and approximately 95% of the target region was covered at least 10×. In total, we detected 32,222 variants: 26,230 single nucleotide polymorphisms (SNPs), 2536 insertions, and 3456 deletions.B3GLCT  that were segregated from an unaffected father and unaffected mother, were strong candidates to be potentially responsible for the observed pathogenicity, which, most likely, was PPS  in the proband. In terms of localization, 38 variants were annotated as exonic and 227 as intronic changes. All discovered variants were concatenated into one VCF file for further analysis . Using i was PPS .B3GALTL, OMIM:610308:0001, RCV000001326.4) associated with PPS. Moreover, using Varsome, the discovered variant was classified according to ACMG2015 as pathogenic, with identified criteria to be PVS1, PP3, PP5.The first variant, inherited from the mother, was a substitution, NM_194318.3:c.660+1G>A (rs80338851), reported as pathogenic allele , which has not been previously reported . In Varsome, the discovered variant was classified according to ACMG2015 as likely pathogenic, with identified criteria to be PVS1 and PM2. Moreover, this mutation had a MutationTaster score of 1 (prediction: Disease causing) and changed the reading frame downstream of the amino acid (AA) at position 252. As a result, the mutation terminated translation at AA position 264, whereas a wild-type protein is 498 AA long. Both mutations were manually reviewed in an integrative genomic viewer (IGV).B3GLCT, we performed Sanger sequencing of exons 8 and 9 in the patient and both parents. Both mutations were present in the patient and the inheritance pattern was confirmed. Sanger sequencing electropherograms are presented in For final confirmation of the potentially causative variant in As reported, 6%–8% of children are born with developmental disorders, with many of them caused by genetic changes in a single gene . DespiteIndependent of the sequencing scale, NGS creates considerable data burden Thus, careful variant filtering and prioritization, according to the information available from sequencing databases, in compliance with ACMG standards are required. Additionally, differences in NGS data analysis between laboratories could generate discrepancies in the diagnosis, thus filtering schemes should be clearly reported. Moreover, clinical interpretation of sequencing data requires matching information on patients’ genotypes and phenotypes and is usually complemented by the integration of information from databases such as OMIM or ClinVar, which are manually constructed and curated. The search for causal variants should be supported by published evidence if it exists, and this requires manual querying of relevant literature databases. This step is the most time-consuming in data processing and has a major impact on timely diagnosis and minimizing false negative and false positive findings. Web-based literature search engines are employed to assist manual curation of newly published information. AMELIE is a method for ranking candidate causal genes related to the phenotype, extracted from primary literature using supervised machine learning methods. The inclusion of HPO-encoded phenotype information facilitates communication with clinical specialists. In our opinion, it combines the convenience of other HPO-based variant ranking tools such as Exomiser  with enrB3GLCT gene, which has been previously associated with the PPS phenotype, observed in our patient. Using only a VCF file, annotated by GEMINI with several pathogenicity prediction tools , resulted in conflicting predictions for variants in some cases. Although the list of possible causative variants included two variants of B3GLCT as compound heterozygous variants, it was not fully informative as the annotation missed the prediction for the deletion NM_194318:c.755delC. Both AMELIE and wANNOVAR ranked variants in genes related to HPO-encoded phenotypes. As a result, B3GLCT got the highest score with both tools. Moreover, in contrast to lower ranked genes, the disease annotated to B3GLCT—PPS-matched our patient’s phenotype perfectly.The combination of these two strategies applied to trio sequencing data led us to the identification of compound heterozygous variants in the B3GLCT mutation was 49 [B3GLCT, encoding an O-fucose-specific β-1,3-glucosyltransferase , responsible for attachment of glucose to O-linked fucose (O-fucose), which is previously added by protein O-fucosyltransferase 2 (POFUT2) to thrombospondin type 1 repeats (TSRs), present in many proteins [POFUT2, B3GLCT provides a noncanonical quality control system of proper protein folding in cells [B3GLCT and ADAMTS9 [PPS, a subtype of ASD, inherited in a recessive manner, is a very rare disease with unknown incidence or prevalence. As reported in 2016 by Jaak Jaeken et al., the worldwide number of patients diagnosed with PPS due to n was 49 . Common n was 49 . In termn was 49 ,23. It iproteins . Localizproteins , beta-1,in cells . A GWAS  ADAMTS9 .B3GLCT gene reported in 10 individuals in the LOVD database. Variant c.660+1G>A (rs80338851), located at the donor splice site (5′ss) of exon 8 observed in our patient, is one of the most common among variants identified in PPS patients [On that date, there were 22 unique variants of the patients . It has patients . The respatients . NMD is patients . In mammpatients .The second of the detected variants, c.755delC (p.Thr252fs), was not present in any publicly available database and is the first one reported in exon 9. Pathogenicity prediction algorithms assigned this variant as disrupting. Similar to c.660+1G>A, c.755delC introduces a PTC within exon 9, 26 nucleotides from the splice site, which also should result in NMD.Our study demonstrates the important role and possible diagnostic utility of NGS combined with AMELIE and wANNOVAR.Such an approach, on the one hand, brings data on genes and variants with well-described roles in disease development based on curated, authoritative databases such as OMIM (wANNOVAR) and, on the other hand, updates the analysis with current information from published manuscripts. Such a procedure helps reduce the number of false positive findings not related to a phenotype which are over-abundant in the VCF file annotated with pathogenicity tools.The patient was recruited from the Clinic of Neonatology at the Jagiellonian University Medical College, Krakow, Poland and the Ophthalmology Clinic and Department of Ophthalmology, Medical University of Silesia in Katowice, Poland. Written informed consent was obtained from the study participants and informed parental consent was obtained on behalf of the child. Developmental and dysmorphology assessments were conducted by a clinical geneticist. This research adhered to the tenets of the Declaration of Helsinki and was performed upon approval of the protocol by the Jagiellonian University Ethics Committee No. 122.6120.12.2015 (29 January 2015).The patient was admitted to the outpatient clinic of the Department of Ophthalmology at the Silesian Medical University in Katowice at the age of seven weeks and then, subsequently, at four and six months.DNA was extracted from the peripheral blood of the affected child and both his healthy parents with the Maxwell 16 Blood DNA purification kit  on the Maxwell 16 device (Promega). Sequencing libraries were prepared with the TruSight one sequencing panel kit  according to the manufacturer’s protocol. In brief, 50 ng of DNA was fragmented and adaptor-tagged in an enzymatic reaction. Genomic libraries were enriched for the coding regions of the 4813 genes by two cycles of hybridization with biotinylated probes, followed by capture on streptavidin beads; 12.5 pM libraries were sequenced on the MiSeq sequencer (Illumina) using v3 chemistry reagents (2 × 150 bp reads).Raw reads were processed with the Illumina software, generating base calls and corresponding base-call quality scores. These data were then processed through our custom pipeline that uses open-source programs . BrieflyB3GLCT (RefSeq NM_194318) were analyzed by Sanger sequencing on the ABI3500 sequencer . The obtained sequences were aligned to the reference NC_000013 with the SeqScape software . After identification of an exon-shortening event, the overlapping sequence resulting from the heterozygous deletion was analyzed manually using SnapGene .For final confirmation of the most plausible causal variants, exons 8 and 9 of


Primary hepatic mucosa-associated lymphoid tissue (MALT) lymphoma is a rare disease, and there is no consensus yet on the treatment modalities. Here, we report a new case of MALT lymphoma and review the current literature on this disease.A 73-year-old man was admitted to our department following the incidental finding of a solitary 1.8-cm diameter mass in the liver.Microscopic findings identified the mass as a tumor with infiltration of diffuse atypical B lymphocytes. Immunohistochemical analysis showed positivity for CD20 and CD79a, and negativity for CD3 and CD5. These collective data led to the diagnosis of primary hepatic MALT lymphoma.The tumor was removed by surgical resection. The patient refused additional treatment after the surgery.At the time of writing this report, the patient has been disease free for 6 months postsurgery.Review of the previously published case reports on this rare tumor type indicates that in addition to chronic liver inflammation due to infection or other reasons, genetic aberrations can also contribute to the development of hepatic MALT lymphoma. Additionally, IgH rearrangement is a good genetic hallmark of this tumor. Owing to no specific clinical or radiologic features to define the disease profile for diagnosis, surgery may be a good choice for both diagnosis and therapy if the patient's condition permits. However, subsequent reports have indicated the involvement of organs such as the stomach, lungs, parotid glands, thyroid, and on very rare occasions the liver.,3 To date, the PubMed database includes only 36 articles  describing primary MALT lymphoma of the liver. Unfortunately, no definitive etiologic profile has been established for this disease, and there is no consensus on the best approach for its management. In this report, we discuss our clinical experience of treating a patient with primary hepatic MALT lymphoma. We also present our findings from a systematic review of the related PubMed literature, discussing the possible treatment modalities for this rare disease.Primary hepatic mucosa-associated lymphoid tissue (MALT) lymphoma is a rare disease. It was 1st described by Isaacson et al23 IU/mL, but was negative for hepatitis C virus and human immunodeficiency virus.A 73-year-old man was admitted to our department following the incidental finding of a focal liver mass in computed tomography (CT) scan. The patient reported no symptoms, other than upper abdominal discomfort. He had no significant medical history and no remarkable family history. Physical examination did not show any abnormalities. Findings for all clinical laboratory tests conducted at our institution, including those for the carcinoembryonic antigen and α-fetoprotein tumor markers, were within the normal range Table . HoweverThe mass in the left hepatic lobe (segment II) was detected on plain CT as a hypodense lesion Fig. A. ContraBased on the clinical and radiologic findings, the lesion was diagnosed as malignant. The patient consented to undergo a laparoscopic left lateral segment liver resection (segments II–III). The gross finding following the resection was a white-colored, 1.8 cm, nodular tumor mass Fig. . HistoloBased on the above-mentioned pathologic features, the patient was diagnosed with primary hepatic MALT lymphoma, specifically a low-grade malignant extranodal marginal zone B-cell lymphoma. The patient refused any additional treatment after the surgery and remained disease free throughout the 6 months of follow-up (at the time of writing of this report).3 Today it is described as an extranodal marginal zone B-cell lymphoma, a type of non-Hodgkin lymphoma. The incidence of MALT lymphoma is rare, representing only 7% to 8% of all non-Hodgkin lymphoma cases. It involves a wide variety of organs, such as the gastrointestinal tract, thyroid, lung, parotid glands, breast, and liver.,3 While the most common primary sites are the stomach, thyroid, and lungs, the incidence of MALT lymphoma in the liver is extremely rare.,3The MALT lymphoma was 1st described by Isaacson and Wright in 1983.,6–35 besides the case we are presenting  OR mucosa-associated lymphoid tissue[Title]) AND ((liver[Title]) OR hepatic[Title])” yielded only 45 potentially relevant articles. After reviewing the titles and abstracts for relevance to primary hepatic MALT lymphoma, we had 36 articles. Exclusion of articles without full-text  and English language translation  led to a final count of 31 articles describing 46 casesNonhepatic MALT lymphoma usually arises in the presence of chronic inflammation, due to either an infectious or autoimmune process. However, the etiology of hepatic MALT lymphoma is still unclear. The 47 cases considered in this paper Table  includedHelicobacter pylori infection, thyroid MALT lymphoma with Hashimoto disease, and thymic MALT lymphoma with Sjögren syndrome. Thus, a plausible hypothesis is that chronic inflammation can stimulate the development of MALT lymphoma, with a nodule-like structure of the ileum being formed and marginal zone cells gradually proliferating under the inflammatory stimuli to ultimately develop into a MALT lymphoma. However, it is important to remember that 45% of the cases from the literature had no concomitant liver diseases, and they cannot be explained by the above theory.Different nonhepatic MALT lymphomas have also been associated with inflammatory conditions, such as gastric MALT lymphoma with  According to our literature review (q27:q32), 4 cases of T(14:18)(q32:q21), and 1 case of Trisomy3/Trisomy18 have been reported in MALT lymphomas. Among these 12 cases, while 4 patients had H pylori infection, 1 had HBV infection, and the remaining 7 patients had no known infections. Thus, besides infections, genetic aberrations are also likely to be involved in the development of hepatic MALT lymphoma, and IgH rearrangement might be a good genetic hallmark of this disease.A tumor is essentially a genetic disease, and 4 chromosomal ectopic recombinations have been identified in MALT lymphomas.ew Table , genetic (64 years) and Dong et al (62 years). The clinical presentations in these cases varied, ranging from nonsymptomatic to end-stage liver disease. However, most of the patients experienced no symptoms and were incidentally identified during clinical examinations or surgical explorations.The 47 total cases of hepatic MALT lymphoma included in this review Table , includeThe majority (72%) of the patients with no symptoms (n = 32) presented with a solitary mass. Of these, 17 patients underwent surgical resection, and 3 each received chemotherapy and radiotherapy. The remaining 9 patients presented with multiple masses, and while 2 of them underwent surgical resection with or without chemotherapy, 1 underwent liver transplantation, 2 received chemotherapy, and the remaining 4 patients did not receive any treatment.,27,28,31 hypodense lesions in another 8 cases (CT findings),,17,30,31 and slightly enhanced in 5 cases (with contrast administration).,20,28,29 The MRI revealed the masses appearing as low density T1-weighted images in 8 cases,,17,27,28 as high-density T2-weighted images in 9 cases,,17,27,28 and with restricted diffusion on diffusion-weighted imaging in 1 case. Our patient also showed these radiologic features on CT and MRI. However, these imaging features are similar to those for other malignant tumors of the liver, such as hepatocellular carcinoma, intrahepatic cholangiocellular carcinoma, and metastatic tumor, making it difficult to differentiate them from hepatic MALT lymphomas.The tumors ranged in size from 0.7 to 9.0 cm in diameter, with an average diameter of 3.5 cm. No distinctive radiologic characteristics were apparent. The hepatic MALT lymphomas appeared to be hypoechoic masses in 8 cases (ultrasound studies),In our present case, the patient was a 73-year-old man with HBV infection, and the tumor was found incidentally during a CT scan for upper abdominal discomfort. The solitary mass detected was 1.8 cm in diameter. The patient underwent surgical resection due to a suspicion of hepatocellular carcinoma, which led to the fortuitous diagnosis of primary hepatic MALT lymphoma. As of when writing this report, the patient has remained disease free without any additional therapy  since the surgical treatment.To date, no optimal therapeutic strategies have been established for the clinical management of primary hepatic MALT lymphomas, because of the small number of patients worldwide. With regards to the indolent and favorable prognosis of this tumor Table , in mostRecurrence was reported in 5 cases  with relapse times ranging from 12 to 96 months. These cases included 1 each involving the pulmonary system, pulmonary system 1st and the parotid gland in a 2nd recurrence, and liver. The involved organs for the remaining 2 cases were not reported. Of these 5 cases, 1 patient refused any further treatment. The patient who experienced 2 relapses received rituximab after the 1st relapse, which resulted in complete remission for 84 months, and then received idelalisib after the second relapse, again achieving complete remission. The remaining 3 patients underwent either resection, chemotherapy or radiotherapy only, and all of them achieved complete remission.Considering the data for the 47 collective cases reported, hepatic MALT lymphoma is an extremely rare disease, tending to be a solitary and small tumor, mostly occurring in elderly people. The etiology of hepatic MALT lymphoma is still unclear, though chronic liver inflammation due to infections or other reasons, and genetic aberrations might be contributory factors to its development. Owing to the general findings of no specific clinical presentation or distinctive radiologic features, it is often difficult to make a definite diagnosis of hepatic MALT lymphoma before tissue resection and histologic confirmation. our review of relevant literature, and on the disease's indolent characteristics, the reasonable choices for its management include surgery, radiotherapy, chemotherapy, targeted therapy alone or in combination, or even simply observation. Since it is challenging to diagnose hepatic MALT lymphoma before the operation and there are concerns regarding needle metastasis of a malignant tumor, surgery is probably the best choice for both diagnosis and therapy, as long as the patient's physical condition permits. However, the treatment choice should be based on the size, location, and number of the tumors as well as the patient's performance status. Additional reports will enhance our knowledge of this rare disease and help improve its management.To date, no standard management modalities have been recommended for this rare disease. Based on the National Comprehensive Cancer Network (commonly known as the “NCCN”) guidelines for non-Hodgkin lymphomas,The authors thank Dr Hua Liu, Jue Wang, and Jing Chen for their technical support and for providing the pathologic pictures.Conceptualization: Jian Lv.Data curation: Yong Ji.Formal analysis: Yong Ji.Supervision: Xin Jian Du, Xin Yang.Writing - Original Draft: Hua Zhi Xie.Writing - Review & Editing: Hua Zhi Xie, Jian Lv.


Mcr, was undetected but more carbon fixation genes than nitrate reduction and sulfate genes were found. This suggests that there may be a considerable number of autotrophic microorganisms that cause the phenomenon of low concentration of dissolved inorganic carbon in contaminated areas. The metagenomics data also revealed that most heterotrophic, sulfate, and nitrate reduction genes in the aquifer were assimilatory sulfate and dissimilatory nitrate reduction genes. Although there was limited dissolved oxygen, aerobic degrading genes AlkB and Cdo were more abundant than anaerobic degrading genes AssA and BssA. The metagenomics information can enrich our microorganic knowledge about petroleum-contaminated aquifers and provide basic data for further bioremediation.In petroleum-contaminated aquifers, biodegradation is always associated with various types of microbial metabolism. It can be classified as autotrophic (such as methanogenic and other carbon fixation) and heterotrophic (such as nitrate/sulfate reduction and hydrocarbon consumption) metabolism. For each metabolic type, there are several key genes encoding the reaction enzymes, which can be identified by metagenomics analysis. Based on this principle, in an abnormally low dissolved inorganic carbon (DIC) petroleum-contaminated aquifer in North China, nine groundwater samples were collected along the groundwater flow, and metagenomics analysis was used to discover biodegradation related metabolism by key genes. The major new finding is that autotrophic metabolism was revealed, and, more usefully, we attempt to explain the reasons for abnormally low DIC. The results show that the methanogenesis gene,  The biodegradation of petroleum plays a vital role in transforming hydrocarbons into harmless matter in the natural environment . The und2) and inorganic oxide, such as nitrate (NO3−), manganese (IV) (Mn4+), ferric ion (Fe3+), sulfate (SO42−), and CO2, are considered to be electron acceptors [It is generally believed that the biodegradation of petroleum hydrocarbons consists of oxidoreductase biochemical reactions caused by microorganisms in aquifers. In the reactions, petroleum hydrocarbons are considered to be electron donors, while dissolved oxygen  Mn+, ferriccceptors .AlkB, AssA, and BssA), methanogenic genes , and carbon fixation genes , as well as key genes of the nitrogen/sulfur cycle  were explored [To reveal such metabolism in PHC aquifers, many studies have been carried out. Most of these studies focused on the relationship between microbial community diversity and environmental factors, and it is generally acknowledged that hydrocarbon biodegradation microorganisms vary depending on sampling depth, contaminant concentration, redox conditions, groundwater flow field conditions, and so on ,7,8. Comexplored ,13,14,15explored  quantitaFittingly, contrary to the general law that dissolved inorganic carbon (DIC) is usually affected by organic carbon consumption metabolism producing DIC, an abnormal phenomenon of low DIC, possibly caused by carbon fixation, was discovered in a PHC aquifer in North China. Biodegradation related metabolism might well explain that abnormal phenomenon. To obtain sufficient and reliable evidence, nine monitoring wells along the groundwater flow were sampled. A metagenomics approach combined with next-generation sequencing was carried out to obtain a broad-spectrum profile of microbial information in collected samples. Functional processes related to energy metabolism, including carbon fixation pathways in prokaryotes; nitrogen, sulfur, and methane metabolism; and hydrocarbon degradation, were analyzed based on the metagenomics data. The results could give us a new knowledge about petroleum-contaminated aquifers and be helpful for the management of these contaminated sites.3−, and SO42−). Therefore, MW14 can be considered as the background well. Along the groundwater flow, MW4 and NA125 were designed as the upstream wells, and PM3 and MW3 wells were designed as the contaminated source wells, while NA68 and MW6 were designed as the downstream wells. PM7 and NA7 were designed as transitional wells to monitor the information of the zone adjacent to the contaminated source zone.The sampling site is located at a former petrochemical plant in the piedmont plain of North China. The site was contaminated decades ago by petroleum leaked from storage tanks. The contaminants on the surface have been removed, but the groundwater at the site is still contaminated by petroleum. The groundwater flow is from northwest to southeast/east. Along the groundwater flow, monitoring wells were designed in and around the leakage zone to monitor the information of contaminated groundwater . There wThe aquifer at the site is mainly composed of sand gravel layers. The thickness of the aquifer is about 3.3 m. Its lower layer is silty clay, which can be considered as the aquiclude. Its upper layer is also silty clay at most of the site, while in the downstream area (around NA68 and MW6) there is no upper aquiclude. The groundwater depth is about 21.7 m to 24.0 m and the water table is higher than the upper confining bed. Therefore, the groundwater is confined in the main area and unconfined in the downstream area. The groundwater flow is from northwest to southeast.2 analysis. The samples were then stored on ice in an incubator before being transported to the laboratory. For the metagenomics analysis, 5 L of groundwater was collected in a sterilized 5 L plastic bucket and stored on ice in an incubator. The water samples were transported to the laboratory, and DNA was collected in five poly tetra fluoroethylene filter membranes with a pore size of 0.22 μm by air pump filtration in one day. The filter membranes were stored at −80 °C until DNA extraction.Samples were collected from the monitoring wells using bailers in August 2018. Information about the water temperature (T), pH, electrical conductivity (EC), dissolved oxygen (DO), and oxidation–reduction potential (ORP) in the collected groundwater was determined using a Portable Parallel Analyzer  and recorded before collecting the laboratory testing samples. The groundwater samples were considered to be representative when the variations of T, pH, EC, DO, and ORP in three successive samples were within ±1 °C, ±0.2, ±3%, ±10%, or ±0.2 mg/L, and ±20 mV, respectively. When sampling, 1 L of groundwater was collected in two 500 mL plastic buckets for inorganic analysis and 160 mL was collected in four 40 mL amber glass bottles with Teflon airtight caps for organic and CO2+, Mg2+, Na+, K+, NO3−, SO42−, HCO3−, Cl−, Fe2+, Mn2+, and CO2, were measured following standard methods [Concentrations of the monocyclic aromatics benzene, toluene, ethylbenzene, m-xylene, p-xylene, and o-xylene (BTEX), as well as volatile organic compounds (VOCs), were determined according to the US Environmental Protection Agency (EPA) Methods 8260 and 8015d ,18. Conc methods .® DNA Kit  according to the manufacturer’s instructions. Concentration and purity of extracted DNA were determined with TBS-380 and NanoDrop2000, respectively. DNA extract quality was checked on 1% agarose gel.Total genomic DNA was extracted from nine collected samples using the E.Z.N.A.TM DNA Sample Prep Kit . Adapters containing the full complement of sequencing primer hybridization sites were ligated to the blunt ends of fragments. Paired-end sequencing was performed on an Illumina HiSeq4000 platform  at Majorbio Bio-Pharm Technology Co., Ltd.  using a HiSeq 4000 PE Cluster Kit and HiSeq 4000 SBS Kit according to the manufacturer’s instructions (www.illumina.com).DNA extract was fragmented to an average size of about 300 bp using Covaris M220  for paired-end library construction. A paired-end library was constructed using a TruSeq(1) Sequence quality control and genome assembly:https://github.com/jstjohn/SeqPrep). Low-quality reads  were removed by Sickle (https://github.com/najoshi/sickle). Metagenomics data were assembled using MEGAHIT (https://github.com/voutcn/megahit), which makes use of succinct de Bruijn graphs. Contigs with a length of 300 bp or more were selected for the final assembling result, and then the contigs were used for further gene prediction and annotation.Adapter sequences were stripped from the 3’ and 5’ ends of paired-end Illumina reads using SeqPrep ((2) Gene prediction, taxonomy, and functional annotation:http://metagene.cb.k.u-tokyo.ac.jp/). The predicted ORFs with a length of 100 bp or more were retrieved and translated into amino acid sequences using the National Center for Biotechnology Information (NCBI) translation table.Open reading frames (ORFs) from each assembled contig were predicted using MetaGene (http://www.bioinformatics.org/cd-hit/), and the longest sequences from each cluster were selected as representative sequences to construct a non-redundant gene catalog. Reads after quality control were mapped to the representative sequences with 95% identity using SOAPaligner (http://soap.genomics.org.cn/), and gene abundance in each sample was evaluated.All predicted genes with a 95% sequence identity (90% coverage) were clustered using CD-HIT  for taxonomic annotation.Representative sequences of the non-redundant gene catalog were aligned to the NCBI NR database with an e-value cutoff of 10Functional read annotations for presented figures and tables were assigned using the Enzyme database of the Kyoto Encyclopedia of Genes and Genomes (KEGG). Gene expression pathways were explored/visualized using the KEGG mapper . Parts pContaminant concentrations, electron acceptors, metabolic byproducts, and other hydrochemical parameters of the collected samples from August 2018 are listed in All the contamination indices indicated that the aquifer was still contaminated by petroleum hydrocarbons. There was no sample free from contamination in the aquifer. The downstream wells, MW3, NA7, NA68, and MW6, had higher concentrations of contaminants. This indicates that contaminants migrated to the downstream aquifer with groundwater flow.42− concentrations and ORP values had similar distribution: Values were higher upstream and decreased with groundwater flow, while Fe2+ and Mn2+ concentrations were just the opposite. This indicates that there was biodegradation of hydrocarbons, as the electron acceptors were consumed [3− should have had similar distribution according to the natural attenuation theory [3− upstream and downstream but less in the contaminated source area. The most likely cause is that there was outside nitrogen input around NA68. The evidence is that there is a sewage treatment plant nearby and the ammonium concentration in NA68 is particularly high (300 mg·L−1). Even though there was nitrogen input, the concentration decreased compared with the upstream wells. It is suggested that there was also nitrate reduction biodegradation. Our previous study revealed that biodegradation in the aquifer in 2016 was mainly caused by nitrate and sulfate reduction [DO and SOconsumed . NO3− shn theory , but it eduction . It was 2), elevating DIC as a result [Dissolved inorganic carbon (DIC) concentration in wells around the contamination source zone  was lower than that in upstream and downstream wells. It is generally acknowledged that heterotrophic microorganisms completely oxidize organic carbon (hydrocarbons) to inorganic carbon  cycle, Wood-Ljungdahl (WL) pathway, dicarboxylate-hydroxybutyrate (DH) cycle, 3-hydroxypropionate (HP) cycle, Arnon-Buchanan (AB) cycle, and hydroxypropionate-hydroxybutyrate (HH) cycle ,31. The 2 to other material in each pathway is shown in The abundance of genes encoding these enzymes is listed in Reyranella, Novosphingobium, Microbacterium, Sphingomonas, Acidovorax, and Bordetella, potentially have the ability to biodegrade hydrocarbons [Another interesting finding is that the carbon fixation genes were more abundant in the contaminated source area, which was attributed to the higher abundance of carbon fixation genes in the WL and HP/HH pathways. The analysis of species structure  showed tocarbons ,38,39,40ocarbons ,42. It iocarbons . TherefoAlthough the mechanism is still unclear and should be studied further, the result explains the phenomenon of low DIC concentrations in the contaminated aquifer. NA125, PM7, and PM3 wells were less contaminated, and they had lower DIC concentration. In particular, the NA125 well had the most abundant carbon fixation genes among upstream wells  that had fewer contaminants, and therefore had the lowest DIC concentration . MW3, NAThe amount of genes encoding carbon fixation was more than 1500 ppm in all samples. Except for MW6, the abundance of carbon fixation genes was greater than that of genes involved in nitrate and sulfate reduction , both of2 to methane, methanol to methane, and acetate to methane [mer, cdh, and mta is ~70, ~20, and ~3 parts per million (ppm), respectively. Therefore, the pathway of CO2 to methylcoenzyme M is dominant. This process mainly occurs around the contaminated source area (PM3 to MW3). It may be because there was less dissolved oxygen (DO) in the area , the lowest concentration of DO (1.02 mg/L), and adequate nitrate (33.21 mg/L) in the NA68 well. Denitrifying bacteria are always considered to be anaerobic heterotrophic microorganisms [In subsequent steps of nitrite reduction, there are three steps in denitrification and only one step in dissimilatory and assimilatory reduction. In denitrification, nitrite is reduced to nitric oxide through nitrite reductase encoded by the rganisms , and theNirBD and NrfAH genes. The abundance of the two genes was about 100 ppm to 300 ppm, which was greater than that of the NirK (NirS) gene in denitrification except in the NA68 sample. There was more abundance in upstream and downstream samples. There was less of the assimilatory reduction gene, NirA, whose abundance was 0–50 ppm.In the dissimilatory reduction process, nitrite is transformed into ammonium directly through nitrite reductase encoded by Based on the above results, it can be speculated that nitrite reduction was mainly caused by dissimilatory reduction and denitrification in the study aquifer. It is reported that denitrifying microorganisms are capable of oxidizing toluene, ethylbenzene, and m-, o-, p-xylenes, and there was substantial investigation into BTEX degradation under denitrification conditions in aquifers ,49. DeniAmoABCD and Hao genes) in the aquifer for the limited DO in groundwater [There were fewer nitrification genes  in some wells with little abundance, except NA7 and MW6 wells located downstream, with the least dissolved oxygen, to which nitrogenase is sensitive [There was also the nitrogen fixation gene  and Sir(F), which account for 150 ppm to 400 ppm. Interestingly, the abundance of genes involved in each step of assimilatory reduction was less in contamination source area samples  than other samples. The lower abundance of Sat and Ask genes in samples from the contamination source area may be caused by the coaction of redox conditions and contaminant concentrations. The Sat gene is also involved in sulfur oxidation  and Sir genes may share the most abundant microorganism, Novosphingobium, and other microorganisms with the Sat gene , and Sir genes.There are four steps in assimilatory reduction. The genes encoding the enzymes that transform sulfate to (adenylyl sulfate) APS and then (3’-phosphoadenylyl sulfate) PAPS are xidation , which wSat gene , which mSat gene, encoding the first step reaction enzyme, is shared with assimilatory reduction. The second step (APS to sulfite) gene, Apr, and third step (sulfite to sulfide) gene, Dsr, had negligible abundance (~10 ppm) compared with assimilatory reduction. The abundance of Apr and Dsr genes was also less in contamination source area samples. Sulfur oxidation microorganisms, such as Thiomargarita and Thiodictyon, and sulfur-reduction microorganisms, such as Desulfovibrio and Desulfobulbus, coexist in the aquifer , followed by enoyl-CoA hydratase (4.2.1.17), anaerobic carbon-monoxide dehydrogenase (EC 1.2.7.4), glutathione transferase (EC 2.5.1.18), and alcohol dehydrogenase (EC 1.1.1.1). These are always considered as essential enzymes related to the degradation of benzoate, nitrotoluene, fatty acid, ketone bodies, and some amino acids ,63,64,65AlkB, EC 1.14.15.3) and catechol dioxygenase genes  implies the degradation of medium-chain (C5–C22) alkanes and aromatic hydrocarbons, respectively, using oxygen as an electron acceptor [AssA, EC 2.3.1.54) and benzylsuccinate synthase gene  imply the degradation of chain alkanes and aromatic hydrocarbons, respectively, in anaerobic conditions [The presence of the alkane 1-monooxygenase gene , and more Cdo genes and fewer AlkB genes in the contamination source samples . The downstream samples (NA68 and MW6) had a medium abundance of AlkB and Cdo genes, while the PM7 sample had the least abundance of both AlkB and Cdo genes. That may be because there was less contamination and relatively adequate oxygen upstream of the aquifer, and Cdo has higher oxygen capture capacity and is more sensitive to the limited oxygen than Alk B. Although there was more contaminant in NA7, NA68, and MW6, the amplification of Cdo gene was restrained by the limited oxygen.There were more BssA genes and fewer AssA genes compared with AlkB and Cdo genes in all samples (AssA genes in MW6, NA7, and PM7 for the less dissolved oxygen in these samples.Although the aquifer was in an almost anaerobic environment, there were fewer anaerobic biodegradation genes. There were no  samples . The mosThe samples collected in different contaminated areas along the groundwater flow revealed some interesting information about microorganism metabolisms. For examples, in contaminated source zone, there were more abundant genes related with carbon fixation, xenobiotic biodegradation, and catechol dioxygenase, but less alkane 1-monooxygenase gene and sulfate reduction genes.Based on the information revealed above, possible mechanisms of biodegradation related metabolism could be deduced as follows: Petroleum hydrocarbons stimulate hydrocarbon biodegradation microorganisms, which take nitrate and sulfate as both electrical acceptors and nutrient elements. During biodegradation, some biodegradation microorganisms, considered to be facultative autotrophic bacteria, can use inorganic carbon and intermediate degradation products of contaminant components as their carbon source. The metabolism can fix the inorganic carbon and result in lower DIC concentration in the contaminated source zone. These processes could not generate methane, since the easily metabolized electrical acceptors, nitrate and sulfate, were adequate and there was no methanogenesis condition in the aquifer. This deductive mechanism is based on DNA information, which is a good indication to know the situation in an aquifer. To gain more information on the real activity of genes and the reactions acting in this context, an analysis of mRNA should be carried out in our future study.In this study, high-throughput sequencing of environmental DNA was used to gain insight into biodegradation related metabolism in a petroleum-contaminated aquifer. The hydrochemical results suggest that a mass of petroleum hydrocarbons was biodegraded by microorganisms through the metabolism of nitrate and sulfate reduction. Metagenomics analysis proved the speculation and gave more detail information: Nitrate and sulfate reduction were dominated by dissimilatory nitrate and assimilatory sulfate reduction, respectively. This indicates that nitrate and sulfate may serve not only as electron acceptors but also as sources for microorganism metabolism. Although the aquifer was in an anaerobic environment, it had a considerable abundance of aerobic biodegradation genes. Accompanied by biodegradation of hydrocarbons, carbon fixation genes thrived, which probably caused the phenomenon of lower dissolved inorganic carbon concentration in the contaminated source area. Interestingly, no methanogenesis genes were detected in the aquifer, indicating that there was less methane effusing into the atmosphere. Considering the great amount of carbon fixation genes in the aquifer, it can be deduced that there would be a carbon sink effect. These interesting findings motivate us to study the mechanism further.


Plexiform neurofibroma is an uncommon variant of NF-1 characterized by deformed masses growing along the length of nerves and affecting surrounding tissues such as skin, muscle, and bone.1 Although rare, abnormal soft-tissue hypertrophy and bone dysplasia can occur in patients with plexiform neurofibroma. This condition is termed elephantiasis neuromatosa (EN).4 Changes in the vascular and lymphatic system, such as enlarged veins,2 dilated lymphatics,3 enlarged lymph nodes,4 and dermal backflow,4 have been observed in limbs enlarged by EN. Although surgery is the standard treatment for plexiform neurofibroma,5 conservative treatment might be an option, especially for more complicated cases such as EN. Here, we describe a custom boot-type brace that we used to treat EN.Neurofibromatosis type 1 (NF-1) is a rare autosomal dominant disease that affects approximately 1 in 3,500 people.A 59-year-old man was referred for evaluation of a giant mass in the right lower extremity from just below the knee to the foot that was diffusely swollen and drooping. He had previously been diagnosed with NF-1 based on clinical symptoms. The circumference of the right lower leg was up to 61.5 cm, compared to 33 cm on the left. The patient was a welder and the mass was frequently injured while walking around at his workplace. Surgery was not indicated because of the risk of injuries to the neurovascular bundles. Because the patient had a history of severe cellulitis and abscess formation , vegetable tanned leather; toe box, cotton and styrene latex resin; heel, rubber and foam rubber; insole core, ethylene vinyl acetate (EVA); and insole cover, lightweight EVA, natural zeolite, and polyester fiber. We thank the patient for granting permission to publish this information.


The onset of most lifetime mental disorders occurs during adolescence, and the years in college, as the final stage of adolescence in a broad sense, deserve attention in this respect. The psychological well-being of undergraduate students can influence not only their academic and professional success, but also the development of society as a whole. Although previous studies suggested psychiatric disorders are common in the adult population, there was little consistent information available about undergraduate students’ mental health problems. This research aimed to describe the changes in depression, anxiety, and stress of Chinese full-time undergraduate students as they experienced university life using the Depression Anxiety Stress Scales-21 (DASS-21). The main conclusions of our study were as follows: (1) on average, students’ severity scores of depression during the four academic years varied between 7.22 and 7.79, while stress scores ranged from 9.53 to 11.68. However, the anxiety scores of college students in the first three years turned out to be 7.40, 7.24 and 7.10, respectively, slightly overtaking the normal threshold of 7. These results indicated that Chinese college students, in general, were mentally healthy with regard to depression and stress, but their average anxiety levels were beyond normal in the first three years. (2) As for the proportions of students with different degrees of severity, approximately 38% to 43% of college students were above the normal level of anxiety, about 35% above the normal level of depression, and around 20% to 30% above the normal level of stress. (3) There were significant differences in the psychological health states of students of different years, especially among the sophomores, juniors, and seniors; the highest score of depression, anxiety, and stress all appeared in the first or second year on average, but some improvements were achieved in the third and last years. The findings suggested that colleges and universities need to pay special attention to psychologically unhealthy students, and with concerted efforts by the government, formulate mental health policies in the prevention, detection, and treatment of students’ psychiatric disorders, rather than just focusing on their average levels of mental health. Mental health serves as an integral component of overall health and well-being, and plays an important role in maintaining physical health. Previous studies also found that adverse physical health status  had an impact on mental health in young people and adults ,2,3,4,5.In the past decades, the number of people suffering from depression and anxiety have increased sharply, which has posed a grave threat to society. Relevant research found that the global aggregate point prevalence of depression was 12.9 percent, by combining the data of over 1 million participants from 30 countries between 1994 and 2014. South America had the highest depression rate, reaching up to 20.6 percent, followed by Asia 16.7%), North America (13.4%), Europe (11.9%), Africa (11.5%), and Australia (7.3%) ,16,17. A.7%, NortCollege is a crucial period of life for students to shape proper values, worldviews, outlooks of life, and resilience . This peMost of the previous studies used cross-sectional data, while some research employed longitudinal designs to study college students’ psychological well-being status, as well as its determinants, in different years. Globally, Puthran et al. (2016) found that medical freshman students had the highest rates of depression at 33.5 percent, which then experienced a significant decline over time to 20.5 percent before graduation . A similIn general, existing literature regarding the psychological distress of college students mainly used cross-sectional data, and the few longitudinal studies often had a small sample size, with participants mostly from a single university. Thus, there is a dearth of detailed analysis and comparisons of the mental health status of college students over the four-year span, especially in a Chinese context. Indeed, college students of different years may be exposed to different learning circumstances and experience various degrees of depression, anxiety, and stress. It is critical for colleges to understand students’ emotional changes in order to offer proper guidance and support. On this base, our study followed a cohort of undergraduates from 15 Chinese universities; using descriptive statistics and multiple group analysis, we pictured the changing trends of students’ negative emotions during their four academic years in college.This study employed data from the “Beijing College Student Panel Survey” (BCSPS) of the “China Education Panel Survey” (CEPS). The sampling frame of this survey was the students’ status data bank provided by the Beijing Municipal Commission of Education. Participants of the survey were randomly chosen from students admitted to 15 universities in the years of 2006 and 2008, and these two cohorts were tracked for four consecutive years from June 2009 to June 2012. The first three rounds were carried out on-site with the cooperation of Beijing Municipal Commission of Education and the Office of Student Affairs of the universities; thereafter, in year four, the participants were invited via text messages and e-mails to log in to the questionnaire website with a unique code, and this round of investigation was conducted entirely online . In ordeIn our study, students’ psychological well-being was assessed using the Depression Anxiety Stress Scales-21 (DASS-21), a self-report measure of the severity of three related negative emotional states. The DASS-21 has been widely recognized for its reliability and different forms of validity in a range of studies from different countries with different samples ,43. EachRegarding the differences in the mental states of college students across years, With the above analysis, we can conclude that Chinese college students were mentally healthy on average, while the psychological health states of freshmen and sophomores warrants more attention. Descriptive statistics showed that students in their first year of college tended to suffer more from anxiety, while in the sophomore year they experienced relatively high levels of stress and depression. As they entered the last two years of college life, their situation seems to have taken a favorable turn. As for the proportions of students with different degrees of mental health problems, approximately 38 to 43 percent of college students had above the normal levels of anxiety, while around 35 percent had above the normal levels of depression. In addition, students whose stress scores surpassed the clinical cutoff values constituted about 20 to 30 percent of the total. The onset of most lifetime mental disorders occurs before young adulthood , and theConsidering the different academic and employment pressures confronted by students of different years in college, there still needs to be a greater exploration than merely descriptive statistics to conclude whether there exist significant differences in the psychological well-being of college students across years. Multiple group analysis is used to determine whether parameters of a measurement model and/or a structural model are invariant across two or more groups . The inv2 (6) = 43.823, p < 0.05; NFI = 0.995; NNFI = 0.991; CFI = 0.995; RMSEA = 0.034). The χ2 difference test between the baseline model and the measurement weights model was significant (χ2 (6) = 43.823, p < 0.05), suggesting that the factor loadings of all year groups were different. In the subsequent model, both factor loadings and variances were constrained equally across years, and the result showed that this was also acceptable (χ2 (9) = 91.346, p < 0.05; NFI = 0.989; NNFI = 0.986; CFI = 0.99; RMSEA = 0.04). The χ2 difference test between these two constrained models was also significant (χ2 (3) = 47.522, p < 0.05). This suggested that, apart from the factor loadings, unique variances of each item were also different in different academic years. Finally, by holding factor loadings, variances and residuals all equal, the multi-sample analysis revealed this constrained model was acceptable (χ2 (18) = 261.342, p < 0.05; NFI = 0.968; NNFI = 0.98; CFI = 0.97; RMSEA = 0.049). The χ2 difference test between the structural covariances model and the measurement residuals complete invariance model was significant (χ2 (9) = 169.996, p < 0.05). Overall, these results indicated that the factor loadings, unique variances, and factor variances were different across years, revealing that there were significant differences in psychological well-being among different years in college. Yet it is still hard to determine which of the four years were different, so we conducted a further comparison between different academic years, as reported in p > 0.05). Nevertheless, the mental health states between the sophomore and junior years differed significantly, with p < 0.05 for the structural covariances model and the measurement residuals complete invariance model. A similar pattern applied to the comparison between the junior and senior years . We also performed difference tests between year 1 and 3, year 1 and 4, and year 2 and 4 for college students as a validation. This indicated that strictly speaking, i.e., by holding factor loadings, variances and residuals as invariant, there were statistically significant differences between the latter three years.The results showed that there was no significant difference in students’ psychological well-being in the freshman and sophomore years, indicated by the measurement weights model, structural covariances model, and measurement residuals complete invariance model (Generally, Chinese college students experienced more mental problems in the freshman and sophomore years. For freshman students, their anxiety may be partly caused by adjustment disorders, especially when students are separated from their parents and friends and have to orient themselves to new environments. Moreover, given the curriculum organization in Chinese universities, which usually set general courses in the first year and introduce more specialized courses from the second year, sophomores may be confronted with intense pressure from study compared to the previous semesters. This may inevitably lead to higher stress and even depression. Our conclusion partly echoed another similar study conducted in the UK, which indicated that once students started university, a great strain was placed on their well-being, with a significant reduction in levels of distress from semester one to semester two being observed in both year one and year three . HoweverIn addition, the changing trend of students’ psychological well-being in China differs from that in the US. In China, the psychological health of freshmen and sophomores deserves more attention. By contrast, in the US, juniors and seniors scored higher on depression, anxiety, and stress scales compared with freshmen, and students’ top three concerns were academic performance, the pressure to succeed and post-graduation plans . A possiPrevious literature on college students’ psychological well-being was generally based on cross-sectional data, so the measurement may not be consistent when comparing students in different years. Our study, using longitudinal data, helps to strengthen the credibility of relevant conclusions. By tracking the same cohort over four years, we confirmed that there were significant differences for students in different years. We attribute the psychological disparity to grade differences, rather than the inapplicability of DASS-21 scale across years, for mainly two reasons. First, the DASS-21 enjoys worldwide recognition in measuring levels of depression, anxiety, and stress, so we assume that it can be applied to examine students in different years ,52,53,54The primary purpose of this study was to examine whether there were differences in the psychological well-being status of Chinese college students as they experienced university life. Through descriptive statistics and multiple group analysis, we can mainly draw the following conclusions.First, Chinese college students were on average mentally healthy with regards to stress and depression, but they suffered from anxiety beyond normal levels in the first three years.Second, approximately 20 to 40 percent of college students suffered from different degrees of depression, anxiety, and stress. This was consistent with some relevant previous research, which concluded that roughly one-fifth of students struggled with different levels of mental health problems.Third, it has been confirmed that there was a disparity in the psychological well-being of students in different years. To be more specific, the highest average scores of depression, anxiety, and stress appeared either in the first or second year. A relatively large ratio of freshmen suffered from anxiety, while, during their sophomore year, students tended to experience more stress and depression. There were significant differences between sophomore, junior and senior students, showing that their mental health situation had improved over time.Based on the results of this study, we suggest that colleges and universities should provide students with tailored psychological guidance, considering that college students in different years may have differentiated psychological well-being status. Universities may offer proper psychological counseling for freshman students in order to relieve their anxiety and pay special attention to improving the psychological well-being of sophomore students. Furthermore, it is worth noting that the overall optimistic situation of the psychological state of colleges students may be due to their weak perception about changes in their external social environment. In the future, we could study college students’ mental changes after entering the labor market for a certain period. The comparison of their psychological situation at work and during college could be explored to offer more implications on the development of psychological well-being counseling programs in college.


The aim of the present study was to assess the efficacy and safety of percutaneous cannulated screw (PCS) implantation assisted by screw view model of navigation (SVMN) to treat femoral neck fracture (FNF).A 42-year-old male patient suffered from a high falling injury, causing pain, swelling, deformity, and limited mobility on his right hip.He was diagnosed with Garden type I of FNF.PCS implantation assisted by SVMN was used to treat fracture of femoral neck in this patient.The follow up lasted for 48 months. A total of 3 screws were inserted into femoral neck, all exhibiting excellent position. The mean screw deviation was 0.43° and 5.73° of femoral neck-shaft and anteversion angle, respectively. The guide wire drilling attempt of each screw was one-time. The fluoroscopic time lasted 6.3 minutes, the Harris hip scores improved from 67 to 88, and the blood loss was 35 mL. It took 11.7 minutes for designing the screws, 13.9 minutes for implanting the guide wires, and 37.3 minutes for placing the screws. No clinical complications were found during 48-month follow-up visit, including head penetration, implant failure, fracture nonunion, and femoral head osteonecrosis.The study revealed that SVMN is conducive to the PCS insertion for FNF. Our lesson is that the FNF must be well reduction before SVMN assisted PCS placement. Percutaneous cannulated screw (PCS) fixation is considered an acceptable method for treating femoral head fractures because of its own advantages, including minimally invasive, short operation time, and good stability. Accurate cannulate screw placement is considered the most vital manipulation to avoid surgical complications.With the aging of the global population, hip fractures will lead to an increase in emergency admission and surgery.,5 Measures to facilitate PCS implantation include computer-assisted surgery,,7 robot-navigated surgery, and guide wire aiming device surgery. Yet, they are accompanied with many complications, for example, femoral head osteonecrosis, fracture nonunion,,11–13 head penetration, and implant failure.Cannulate screw placement by freehand for treating femoral neck fracture (FNF) obtained outcomes that were considered satisfactory. However, the complication rate of freehand cannulate screw placement reaches 12.5% to 55%.To reduce the incidence of clinical complications, we applied a screw view model of navigation (SVMN) technology in the operation of non-displaced FNF. In this screw view model, the navigation monitor displays the static axial, sagittal, and coronal three dimensional computed tomography (3D CT) of FNF with designed screws, which makes the manipulation simple and accurate. To the best of our knowledge, the application of this novel technology with PCS placement has been rarely reported.2The Second Hospital of Jilin University, Changchun, China, approved the study and the institutional guidelines for the care and treatment of patients were rigorously followed. Informed written consent was obtained from the patient for publication of this case report and accompanying images. (2019) Research and Inspection No. (007).33.1A 42-year-old male patient suffered a high falling injury Table , causing it was classified as Garden I.The hip x-ray exhibited the discontinuity of bone in the right femoral neck Fig. . Three-dThe whole hip was scanned by CT preoperatively, and the result was shown in Fig. 3.2The operation was performed under general anesthesia . Short-acting muscle relaxants were provided only during the intubation. When the anesthesia worked, the patient was placed in the supine position.First, a patient tracker , operated with the Navigation System II-CART II with SpineMap 3D 2.0 software , was outfitted on the iliac crest at the beginning of the operation. The system's C-arm tracker, patient tracker, and guide needle sleeve tracker were all activated. After 190° scanning was performed at the center of the femoral neck, 3D images of the lesion were captured. Subsequently, preoperative and intraoperative CT images were matched to provide clear guidance for the guide needle sleeve.Second, the screw view mode was selected in the navigation workstation. Besides, the position of the guide needle sleeve device was adjusted until the direction of guide needle sleeve completely complied with the planning PCS trajectory, and the guide wire was placed once the right lower corner image showed green Fig. . At the Finally, x-ray and CT scan were performed to verify whether the screw was in good position Figs.  and 5. P3.33.3.1 , grade 1 as good (>5 mm), grade 2 as general (<2 mm), and grade 3 as bad  Table , Table 3rew Fig. , Table 34The postoperative 3D CT shows that the 3 implanted screws were all in position of grade 0. The follow up visit lasted for 48 months. The mean angle of screw deviation was 0.43° and 5.73°, respectively, in the femoral neck-shaft angle, anteversion angle, and details are listed in Table It took 11.7 minutes for designing the screws, 13.9 minutes for implanting the guide wires, and 37.3 minutes for placing the screws. The frequency of drilling attempts of each screw was one-time, intraoperative blood loss was 35 mL, the fluoroscopic time lasted 6.3 minutes and Harris hip scores improved from 67 to 88. No clinical complications, such as fracture nonunion, femoral head osteonecrosis, head penetration, and implant failure, were found during 48-month follow-up Table .5 First, complete an entry point for the 3 screws above the distal end of the lesser trochanter. Second, avoid femoral head penetration. Third, achieve the maximal spread of the 3 parallel screws in the femoral neck. However, it is hard to insert the guide needle at the perfect position. Accordingly, complications  may occur during the operation.,10,12–14 Accordingly, appropriate placement of guide needles is vital to treat FNF with PCS. The use of a SVMN technique was explored to assist in a PCS insertion.Perfect cannulate screw placement should satisfy the following requirements.,19 The navigation of the guide needle's direction in the femoral neck can be clearly seen through the image intraoperatively. As a result, the 3 cannulate screws achieved better parallelism and spread with fewer failed attempts. In the present study, the position of the 3 screws were all classified as grade 0 exhibiting small deviation angle, good spread, and parallelism. Each screw was successfully inserted at its first attempt. Compared with the results of previous findings, the drill attempts here were less, and the fluoroscopy time was shorter. No surgical complication was shown here on follow-up, such as no head penetration and implant failure. This positive result was attributed to the use of the navigation system for preoperative planning and the guidance of PCS insertion with a SVMN.The plan function of computer navigation system help the surgeon calculate the length, angle, entry point, and plan trajectory of the screw preoperatively, and SVMN exhibits a perfect screw position on the static axial, sagittal, and coronal image of 3D CT.,20–22 and femoral head osteonecrosis was up to 17% to 45% with PCS.,20,22,23 The medial femoral circumflex artery and lateral femoral circumflex artery provide 90% of the blood supply to the femoral head. Fractures of the femoral neck can cause arterial damage, thereby leading to avascular necrosis of the femoral head. Rahman et al reported that the risk of avascular necrosis was associated with the degree of displacement. In the present study, patients with Garden type I of FNF found no nonunion and femoral head osteonecrosis during follow-up. The reasons for this good result were concluded. On the one hand, the medial and lateral femoral circumflex arteries were not seriously damaged since the displacement of the fracture is small. On the other hand, this was because accurate screw placement avoids iatrogenic vascular injury and stable internal fixation.Fracture nonunion and femoral head osteonecrosis are common complications after FNF surgery with PCS. Previous literatures showed that the incidence of fracture nonunion was ranged from 10% to 47%,Our SVMN technology has the following advantages in design: first, it avoids the effects of multiple drilling on blood supply of femoral head. Second, intraoperative fluoroscopy time was reduced, and radiation exposure of patients and medical staff was reduced. Third, accurate placement of cannulated screw  cannot only increase the stability of fracture fixation, but also avoid screw cutting bone cortex.For the drawbacks of this technology, it has narrower range of indications in the navigation system, it is only suitable for minimal dislocation of FNF, for example, Garden type I and Garden type II, and Garden Type III and Garden IV of FNF require good reduction before SVMN guided PCS fixation. Furthermore, the sample size here is very small, and more samples are required to assess the safety and effectiveness of this technology.In conclusion, the PCS insertion assisted by SVMN technology is a simple and highly feasible procedure. Our lesson is that the FNF must be well reduction before SVMN assisted PCS placement.The authors gratefully acknowledge the cooperation of the doctors and nurses in the operating room.Conceptualization: Bao-Ming Yuan, Dong-Sheng Wang.Data curation: Yi-Kun Jiang.Investigation: Rong-Peng Dong.Methodology: Xin Zhao.Project administration: Dong-Sheng Wang, Ming-Yang Kang.Supervision: Jianwu Zhao.Validation: Yi-Kun Jiang.Writing – original draft: Tong Yu, Bao-Ming Yuan, Xi-Wen Zhang, Rong-Peng Dong, Xin Zhao, Jianwu Zhao.Writing – review & editing: Tong Yu.


Essential hypertension is one of the most common chronic diseases in the world and a major risk factor for cardiovascular and cerebrovascular diseases. Hypertension often leads to a variety of complications, of which vascular endothelial dysfunction is an important part. Traditional Chinese medicine (TCM) combined with western medicine can significantly improve vascular endothelial function in patients with hypertension, but it has not been systematically evaluated for efficacy and safety of essential hypertension. Therefore, we aim to conduct a systematic review and meta-analysis to evaluate the efficacy and safety of TCM combined with western medicine in improving vascular endothelial function in patients with essential hypertension.We will search PubMed, Cochrane Central Register of Controlled Trials (CENTRAL), Excerpta Medica Database (EMBASE), China National Knowledge Infrastructure Database (CNKI), Wanfang Database, China Science Journal Database (VIP Database) and China Biomedical Literature Database (CBM). Clinical trial registrations, potential grey literature, related conference abstracts, and reference lists of identified studies will also be retrieved. The electronic database will be searched for literatures published from the beginning to October 2018. Based on the heterogeneity test, data integration is performed using a fixed effect model or a random effects model. Changes in blood pressure and endothelial function will be assessed as primary outcomes. Drug use, disease progression and adverse events will be assessed as secondary outcomes. RevMan V.5.3.5 will be used for meta-analysis.This systematic review and meta-analysis will provide high-quality evidence from a variety of aspects, including efficacy, blood pressure, vascular endothelial function and adverse reactions, to assess the efficacy and safety of TCM combined with western medicine in patients with hypertension.This systematic review will determine whether TCM combined with western medicine provides evidence for effective intervention of vascular endothelial function in patients with essential hypertension.This systematic review and meta-analysis of randomized controlled trials does not require ethical recognition, and the results of this paper will be published in an open access, internationally influential academic journal.CRD42019140743 The primary goal of hypertension treatment is to reduce the overall risk of heart, brain, kidney and vascular complications development, and death. Common antihypertensive drugs include calcium channel blockers, angiotensin converting enzyme inhibitors, angiotensin receptor blockers, diuretics, and β-blockers. However, due to cost, adverse reactions, complications, etc, nearly half of patients are unable to effectively control blood pressure through drugs.Essential hypertension is the most common chronic disease and a major risk factor for cardiovascular disease. The main complications include stroke, myocardial infarction, heart failure, and chronic kidney disease, which not only lead to high disability and mortality, but also require a large amount of health care and social resources. Hypertension has caused a heavy burden on families and national health care, and has become an important public health problem in China. A large number of studies have shown that TCM decoction, Tai Chi, and acupuncture can effectively treat hypertension.–11 Nowadays, the treatment of hypertension with TCM combined with western medicine is more and more concerned by clinical doctors and patients, and improvement of the vascular endothelial function of hypertensive patients can improve its prognosis. There is no clinical evidence for the improvement of vascular endothelial function in patients with hypertension in comparison with TCM and western medicine. Therefore, we have developed a protocol for systematic evaluation and meta-analysis to evaluate the efficacy and safety of TCM combined with western medicine in improving vascular endothelium in patients with hypertension, and can provide reference applications for clinical use.Traditional Chinese medicine (TCM) is an important part of complementary and alternative medicine. Although the mechanism is not clear, it has been widely accepted in China and applied in clinical practice.22.1The protocol of this review has been registered with the international Prospective Register of Systematic Reviews  and has been reported in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-analyses guidelines.2.22.2.1Inclusion criteria: Randomized controlled trials (RCTs) that use Chinese herbal medicine (CHM) combined with western medicine to treat hypertensive patients, regardless of blinding, will be included in this study.Exclusion criteria: non-RCTs; participants do not meet the inclusion criteria; studies which have been published repeatedly; therapeutic measures failing to meet the predetermined inclusion criteria; control group not western medicine; (6) no blood pressure data for extraction. Language and time will not be restricted to minimize publication bias.2.2.2Inclusion criteria: patients aged 18 and over, but under 80 years of age; patients with a diagnosis of hypertension that is consistent with past guideline definitions (SBP ≥ 140 mm Hg or DBP ≥ 90 mm Hg).Exclusion criteria: Subjects with hypertension and other serious cardiovascular diseases, hepatic failure, and renal failure; subjects with secondary hypertension; subjects with gestational hypertension; and subjects with isolated systolic hypertension.2.2.3This study focuses on the RCTs of hypertension with the therapy of CHM combined with western medicine, and the results will provide advice and consultation for clinicians. All types of CHM, combined with western medicine, with no limitations on the dose, the method of dosing, the composition of the formula, or the duration of administration. Studies that with combination therapy fail to objectively evaluate the efficacy and safety of CHM will be excluded. Studies of control groups were treated with western medicine.2.2.4All types of CHM, combined with western medicine, with no limitations on the dose, the method of dosing, the composition of the formula, or the duration of administration.2.2.4.1: markedly effective = diastolic blood pressure decreased >20 mm Hg (1 mm Hg = 0.133 kPa) or decreased <10 mm Hg, but has reached the normal range of blood pressure; effective = diastolic blood pressure decreased >10 to 19 mm Hg or decreased <10 mm Hg, but has not reached the normal range the normal range of blood pressure; invalid = did not meet the above criteria. Meanwhile, nitric oxide, endothelin-1, flow-mediated dilation, vascular endothelial growth factor, high-sensitive C-reactive protein, angiotensin II, von Willebrand factor, and transforming growth factor β-1 are also included.The primary outcomes will include SBP, DBP, 24 hour SBP, and 24 hour DBP. The therapeutic effectiveness is determined according to the 2010 Chinese guidelines for the management of hypertension2.2.4.2The secondary outcomes are any adverse events .2.32.3.1Seven electronic databases including: PubMed, Embase, CENTRAL, CBM, CNKI, Wanfang Data, and VIP database. These databases were searched from inception to April 2019 for the relevant RCTs of CHM for essential hypertension. Search terms to be used will include essential hypertension, CHM, and RCTs. The search strategy for Medline will be searched via PubMed and is shown in Table 2.3.2A reference list of research and systematic reviews will be reviewed and retrieved for additional testing. Potential gray literature will be searched in OpenGrey.eu. We will search for relevant conference abstracts to find eligible trials. In addition, we will search the World Health Organization International Clinical Trial Registration Platform (ICTRP) and the clinical trial website ClinicalTrials.gov for all new comments related to this topic.2.3.32.3.3.1According to the Cochrane Collaboration System Evaluator's Manual 5.1.0), the basic process of the included literatures will be determined. An excel spreadsheet will be used to record the study name, author, year, database, and whether the study met eligibility criteria and should be included in the review. During abstract screening and full-text evaluation, reasons for inclusion and exclusion are also recorded in a spreadsheet. The records in this spreadsheet will be used to generate the Preferred Reporting Items for Systematic Reviews and Meta-Analyses flowchart  or standard mean difference (SMD) with 95% confidence intervals (CIs), and the dichotomous outcomes will be estimated by the risk ratio (RR) with 95% CIs.2.3.3.5If the required data is unclear or not reported in the clinical paper, the reviewer will contact the 1st author or corresponding author of the study by phone, email, or mail to collect the missing data in the data collection form.2.3.3.6a = 0.1), the value of which is determined by I2. I2 < 50% will be considered to contain small heterogeneity of included studies, while I2 > 50% will be considered to have greater heterogeneity. We will select subgroup analysis and sensitivity analysis to detect sources of possible clinical or methodologic heterogeneity.The heterogeneity of the included studies will be analyzed by Chi-squared test 2.3.3.10If sufficient studies are included, we will conduct a sensitivity analysis to test the robustness and reliability of the results. Sensitivity analysis focuses on the characteristics or types of research, such as methodologic quality, and examines the effects of overall outcomes by excluding certain low-quality studies or nonblind studies.2.3.3.11The GRADE profiler software  will be used to analyze the quality level of evidence.2.3.3.12Since our research does not include information identifying individuals, there is no need for ethics approval. The study will be published in peer-reviewed journals or related conferences.3 Antihypertensive drugs are the main method for the treatment of hypertension, but due to various limitations such as adverse reactions and complications in clinical use, it is difficult to achieve the desired therapeutic goals. Interventions such as lifestyle behaviors and diet control can be used to lower blood pressure, but these interventions are difficult to adhere to. Endothelial injury caused by hypertension has received increasing attention. TCM has been used as an alternative therapy. Many studies have shown that TCM can effectively lower blood pressure and improve vascular endothelial function in patients with hypertension, reducing the occurrence of complications.,17 However, there is no systematic review and meta-analysis of its impact and safety. Therefore, it is necessary to conduct high-quality systematic reviews and meta-analysis. We hope that this systematic review will provide more persuasive evidence for the treatment of essential hypertension with TCM and western medicine. However, there may be some potential shortcomings in the systematic review. First, heterogeneity risks can occur due to different doses, age and severity of hypertension. Second, a small sample may lead to a high risk of bias.Hypertension is the most common chronic disease in the clinic and is an important risk factor for cardiovascular and cerebrovascular diseases.Conceptualization: Weiquan Ren, Li Huang.Data curation: Weiquan Ren, Jiangquan Liao, Jialiang Chen.Formal analysis: Weiquan Ren, Jiangquan Liao, Jialiang Chen.Project administration: Weiquan Ren, Li Huang.Supervision: Li Huang, Zhonghao Li.Writing – original draft: Weiquan Ren.Writing – review & editing: Weiquan Ren, Jiangquan Liao.