Genetic Polymorphisms of Pneumocystis jirovecii in HIV-Positive and HIV-Negative Patients in Northern China.

Pneumocystis jirovecii is an opportunistic fungus that can cause severe and potentially fatal Pneumocystis pneumonia (PCP) in immunodeficient patients. In this study, we investigated the genetic polymorphisms of P. jirovecii at eight different loci, including six nuclear genes (ITS, 26S rRNA, sod, dhps, dhfr and ß-Tub) and two mitochondrial genes (mtLSU-rRNA and cyb) in three PCP cases, including two patients with HIV infection and one without HIV infection in Shanxi Province, P.R. China. The gene targets were amplified by PCR followed by sequencing of plasmid clones. The HIV-negative patient showed a coinfection with two genotypes of P. jirovecii at six of the eight loci sequenced. Of the two HIV-positive patients, one showed a coinfection with two genotypes of P. jirovecii at the same two of the six loci as in the HIV-negative patient, while the other showed a single infection at all eight loci sequenced. None of the three drug target genes (dhfr, dhps and cyb) showed mutations known to be potentially associated with drug resistance. This is the first report of genetic polymorphisms of P. jirovecii in PCP patients in Shanxi Province, China. Our findings expand our understanding of the genetic diversity of P. jirovecii in China.

Clinical features and ultrasound findings of a rare musculoskeletal system disease-neuromuscular choristoma.

BACKGROUND: Neuromuscular choristomas (NMCs), are extremely rare developmental lesions that, have been previously established associated with recurrent fibromatosis after surgery, leading to several operations or even amputation. However, reports on the ultrasound imaging features and clinical conditions of NMCs are rare. The purpose of this study is to describe the ultrasound features and clinical analysis of NMCs to provide suggestions to identify the optimal management strategy. METHODS: From September 2020 to September 2021, 7 patients with a confirmed diagnosis of NMC who underwent ultrasound examination in our department were enrolled in our study. Physical examinations were performed to detect motor deficits, sensory deficits, neuropathic pain, limb undergrowth, muscular atrophy, cavus foot and bone dysplasia. Ultrasound imaging was performed and investigated both in affected nerves and neuromuscular choristomas associated desmoid-type fibromatosis (NMC-DTF). All patients had a definite history and regular follow-up. The clinical course, physical examinations, ultrasound features and pathologic results of NMC patients were analyzed. RESULTS: Seven patients with an average age of 7.0 ± 7.2 years (range: 2-22 years) were enrolled in our study. The affected nerves included the sciatic nerve (6 cases) and the brachial plexus (1 case). Six patients (85.7%) presented with limb undergrowth, 6 (85.7%) with muscular atrophy, and 5 (71.4%) with cavus foot deformity. Based on ultrasound findings, all the visibly affected nerve segments presented with hypoechoic and fusiform enlargement with intraneural skeletal muscle elements. Five patients (71.4%) had NMC-DTFs at the site of the affected nerve. All NMC-DTFs were shown as hypoechoic solid lesions adjacent to the nerve and were well circumscribed. In the subset of the surgery group, all 5 patients presented with progression to NMC-DTFs at the site of the NMCs. No fibromatosis was detected in the other two nonsurgical patients. CONCLUSIONS: Understanding the typical ultrasound features and clinically associated conditions would support the early diagnosis of this rare disease. When a potential diagnosis is determined, an invasive procedure such as biopsy or resection might not be a good choice given the frequent occurrence of complications such as aggressive recurrence.

Direct Repair of the Lower Trunk to Residual Nerve Roots for Restoration of Finger Flexion After Total Brachial Plexus Injury.

PURPOSE: Residual nerve root stumps have been used to neurotize the median nerve in an attempt to restore finger flexion function in patients suffering from total brachial plexus injury. However, the results have been unsatisfactory mainly because of the need to use a long nerve graft. The authors have tried to improve the quality of restored finger flexion by direct approximation of available (ruptured) ipsilateral root stumps to the lower trunk (LT). We sought to validate these results using objective outcome measures. METHODS: This is a study of 27 cases of total posttraumatic brachial plexus palsies. In each case, the neck was explored and ruptured root stumps identified. The LT was mobilized by separating it from the posterior division and the medial cutaneous nerve of the forearm distally. The mobilized LT was then approximated directly to an ipsilateral root stump. The arm was immobilized against the trunk for 2 months. The patients were observed for return of function in the paralyzed upper limb. The presence and strength of finger flexion was measured using the British Medical Council grading. RESULTS: The follow-up period was 36 to 74 months (average, 56.9 ± 13.7 months). Recovery of active finger flexion was M4 in 10 patients, M3 in 8 patients, and M2 to M0 in 9 patients. Meaningful recovery (M3 or greater) of finger flexion was achieved in 18 of 27 patients. CONCLUSIONS: The results of active finger flexion can be improved by direct approximation of the LT to an ipsilateral root stump. TYPE OF STUDY/LEVEL OF EVIDENCE: Therapeutic IV.

Mesenchymal stem cells and mesenchymal stem cell-derived extracellular vesicles: Potential roles in rheumatic diseases.

BACKGROUND: Mesenchymal stem cells (MSCs) have been widely investigated in rheumatic disease due to their immunomodulatory and regenerative properties. Recently, mounting studies have implicated the therapeutic potency of MSCs mostly due to the bioactive factors they produce. Extracellular vesicles (EVs) derived from MSCs have been identified as a promising cell-free therapy due to low immunogenicity. Rheumatic disease, primarily including rheumatoid arthritis and osteoarthritis, is a group of diseases in which immune dysregulation and chronic progressive inflammation lead to irreversible joint damage. Targeting MSCs and MSC-derived EVs may be a more effective and promising therapeutic strategy for rheumatic diseases. AIM: To evaluate the potential therapeutic effectiveness of MSCs and EVs generated from MSCs in rheumatic diseases. METHODS: PubMed was searched for the relevant literature using corresponding search terms alone or in combination. Papers published in English language from January 1999 to February 2020 were considered. Preliminary screening of papers concerning analysis of "immunomodulatory function" or "regenerative function" by scrutinizing the titles and abstracts of the literature, excluded the papers not related to the subject of the article. Some other related studies were obtained by manually retrieving the reference lists of papers that comply with the selection criteria, and these studies were screened to meet the final selection and exclusion criteria. RESULTS: Eighty-six papers were ultimately selected for analysis. After analysis of the literature, it was found that both MSCs and EVs generated from MSCs have great potential in multiple rheumatic diseases, such as rheumatoid arthritis and osteoarthritis, in repair and regeneration of tissues, inhibition of inflammatory response, and regulation of body immunity via promoting chondrogenesis, regulating innate and adaptive immune cells, and regulating the secretion of inflammatory factors. But EVs from MSCs exhibit much more advantages over MSCs, which may represent another promising cell-free restorative strategy. Targeting MSCs and MSC-derived EVs may be a more efficient treatment for patients with rheumatic diseases. CONCLUSION: The enormous potential of MSCs and EVs from MSCs in immunomodulation and tissue regeneration offers a new idea for the treatment of rheumatism. However, more in-depth exploration is needed before their clinical application.

C5aR deficiency attenuates the breast cancer development via the p38/p21 axis.

Emerging evidence has shown activation of the complement component C5 to C5a in cancer tissues and C5aR expression in breast cancer cells relates to the tumor development and poor prognosis, suggesting the involvement of complement C5a/C5aR pathway in the breast cancer pathogenesis. In this study, we found that as compared to the non-tumoral tissues, both C5aR and MAPK/p38 showed an elevated expression, but p21/p-p21 showed lower expression, in the tumoral tissues of breast cancer patients. Mice deficient in C5aR or mice treated with the C5aR antagonist exhibited attenuation of breast cancer growth and reduction in the p38/p-p38 expression, but increase in p21/p-p21 expression, in the tumor tissues. Pre-treatment of the breast cancer cells with recombinant C5a resulted in reduced p21 expression, and MAPK/p38 inhibitors prevented C5a-induced reduction in p21 expression, suggesting the involvement of the MAPK/p38 signaling pathway in the C5a/C5aR-mediated suppression of p21/p-p21 expression. These results provide evidence that breast cancer development may rely on C5a/C5aR interaction, for which MAPK/p38 pathway participate in down-regulating the p21 expression. Inhibition of C5a/C5aR pathway is expected to be helpful for the treatment of patients with breast cancer.

[Tolerance and vegetation restoration prospect of seedlings of five oak species for Pb/Zn mine tailing]. / 5ç§æ Žæ ‘å¹¼è‹—å¯¹é“ é”Œå°¾çŸ¿ç ‚çš„è€æ€§ä¸Žæ¤è¢«æ¢å¤å‰æ™¯.

A pot experiment was conducted to evaluate the growth response and vegetation restoration prospect of seedlings of five oak species for the phytoremediation of lead/zinc (Pb/Zn) mine tailings. Seedlings of Quercus imbricaria, Q. coccinea, Q. pagoda, Q. shumardii, Q. fabri were transplanted into pots containing Pb/Zn mine tailings to comparatively examine their biomass, root morphology, absorption and transfer characteristics of nutrient elements and heavy metals 30 months later. The results showed that all the seedlings could survive in the Pb/Zn tailings after 30 months. The biomass of Q. coccinea and Q. fabri decreased in Pb/Zn tailings compared with the control, while no significant difference were found for other three species. Compared with the control, root biomass was increased to some extent in Pb/Zn tailings except Q. coccinea. The lateral root morphological parameters were reduced only for Q. coccinea . Under heavy metal stress, nutrient concentrations of root and stem of oak seedlings did not change compared with the control. Generally, the concentrations of heavy metals in plant tissues were low, and the values of bioconcentration factor (BCF) and translocation factor (TF) were less than 1. Q. pagoda could accumulate more Cd, with concentrations of 22.4 and 15.1 mg·kg-1 in leaf and stem, respectively, and could translocate more Cd from root to shoot with TF of 2.3. Our results suggested that the seedlings of tested oak species could be used as the potential species for contaminated soil. Q. shumardii had the highest tole-rance with a low BCF and TF, implying that they were better potential candidates for afforestation and ecological restoration of mine tailings.

NKG2A is a NK cell exhaustion checkpoint for HCV persistence.

Exhaustion of cytotoxic effector natural killer (NK) and CD8+ T cells have important functions in the establishment of persistent viral infections, but how exhaustion is induced during chronic hepatitis C virus (HCV) infection remains poorly defined. Here we show, using the humanized C/OTg mice permissive for persistent HCV infection, that NK and CD8+ T cells become sequentially exhausted shortly after their transient hepatic infiltration and activation in acute HCV infection. HCV infection upregulates Qa-1 expression in hepatocytes, which ligates NKG2A to induce NK cell exhaustion. Antibodies targeting NKG2A or Qa-1 prevents NK exhaustion and promotes NK-dependent HCV clearance. Moreover, reactivated NK cells provide sufficient IFN-γ that helps rejuvenate polyclonal HCV CD8+ T cell response and clearance of HCV. Our data thus show that NKG2A serves as a critical checkpoint for HCV-induced NK exhaustion, and that NKG2A blockade sequentially boosts interdependent NK and CD8+ T cell functions to prevent persistent HCV infection.

Excitation Energy-Transfer Processes in the Sensitization Luminescence of Europium in a Highly Luminescent Complex.

The excitation energy transfer (EET) pathways in the sensitization luminescence of EuIII and the excitation energy migration between the different ligands in [Eu(fod)3dpbt] [where fod=6,6,7,7,8,8,8-heptafluoro-2,2-dimethyl-3,5-octanedione and dpbt=2-(N,N-diethylanilin-4-yl)-4,6-bis(3,5-dimethylpyrazol-1-yl)-1,3,5-triazine], exhibiting well-separated fluorescence excitation and phosphorescence bands of the different ligands, were investigated by using time-resolved luminescence spectroscopy for the first time. The data clearly revealed that upon the excitation of dpbt, the sensitization luminescence of EuIII in [Eu(fod)3dpbt] was dominated by the singlet EET pathway, whereas the triplet EET pathway involving T1(dpbt) was inefficient. The energy migration from T1(dpbt) to T1(fod) in [Eu(fod)3dpbt] was not observed. Moreover, upon the excitation of fod, a singlet EET pathway for the sensitization of EuIII luminescence, including the energy migration from S1(fod) to S1(dpbt) was revealed, in addition to the triplet EET pathway involving T1(fod). Under the excitation of dpbt at 410 nm, [Eu(fod)3dpbt] exhibited an absolute quantum yield for EuIII luminescence of 0.59 at 298 K. This work provides a solid and elegant example for the concept that singlet EET pathway could dominate the sensitization luminescence of EuIII in some complexes.

Prognostic value of fibrinogen and D-dimer-fibrinogen ratio in resectable gastrointestinal stromal tumors.

AIM: To investigate the prognostic value of preoperative fibrinogen concentration (FIB) and D-dimer-fibrinogen ratio (DFR) in gastrointestinal stromal tumors (GISTs). METHODS: The purpose of this study was to retrospectively analyze 170 patients with GISTs who were admitted to our hospital from January 2010 to December 2015. The optimal cutoff values of related parameters were estimated by receiver operating characteristic (ROC) curve analysis. The recurrence free survival (RFS) rate was evaluated using Kaplan-Meier curves. Univariate analysis and multivariate Cox regression models were used to analyze the prognostic factors of GISTs. The relationship between the FIB, D-dimer, DFR, platelet count (PLT), and the clinicopathological features of GISTs was described by the chi-square test or nonparametric rank sum test (Mann-Whitney test). RESULTS: In ROC analysis, the optimal cutoff values of FIB, D-dimer, DFR, and PLT were 3.24 g/L, 1.24 mg/L, 0.354, and 197.5 (× 109/L), respectively. Univariate analysis and the Kaplan-Meier survival curve showed that FIB, D-dimer, DFR, PLT, National Institutes of Health (NIH) risk category, tumor size, tumor location, and mitotic index were significantly relevant to the 3-year and 5-year survival rate of patients (P < 0.05). Cox multivariate regression analysis illustrated that FIB (RR: 0.108, 95%CI: 0.031-0.373), DFR (RR: 0.319, 95%CI: 0.131-0.777), and NIH risk category (RR: 0.166, 95%CI: 0.047-0.589) were independent prognostic factors of the RFS rate (P < 0. 05). Moreover, FIB, D-dimer, DFR, and PLT were correlated with the clinical features of GISTs. CONCLUSION: FIB, D-dimer, DFR, and PLT are all related to the prognosis of GISTs. Moreover, FIB and DFR may be independent risk factors for predicting the prognosis of resectable GISTs.

Restoration of active pick-up function in patients with total brachial plexus avulsion injuries.

We designed multiple nerve transfers in one surgery to restore active pick-up function in patients with total brachial plexus avulsion injuries. Forty patients with total brachial plexus avulsion injuries first underwent multiple nerve transfers. These included transfer of the accessory nerve onto the suprascapular nerve to recover shoulder abduction, contralateral C7 nerve onto the lower trunk via the modified prespinal route with direct coaptation to restore lower trunk function and onto the musculocutaneous nerve with interpositional bridging by medial antebrachial cutaneous nerve arising from lower trunk to restore elbow flexion, and the phrenic nerve onto the posterior division of lower trunk to recover elbow and finger extension. At least three years after surgery, the patients who had a meaningful recovery were selected to perform secondary reconstruction to restore active pick-up function. Active pick-up function was successfully restored in ten patients after they underwent multiple nerve transfers combined with additional secondary functional hand reconstructions. LEVEL OF EVIDENCE: IV.

Contralateral C7 Nerve Root Transfer for Function Recovery in Adults: A Meta-analysis.

BACKGROUND: Root avulsion to all 5 roots of the brachial plexus is a common presentation and keeps a major reconstructive challenge. The contralateral C7 (CC7) nerve transfer has been used in treating brachial plexus avulsion injury (BPAI) since 1986. However, the effectiveness of the procedure remains a subject of controversy. The aim of this meta-analysis was to study surgical outcomes regarding motor and sensory recovery after CC7 nerve transfer. METHODS: Chinese or English (i.e., "contralateral c-7", "contralateral c7", "c7 nerve root", and "seventh cervical nerve root") keywords were used for a literature search for articles related to CC7 nerve transfer in several databases (i.e., PubMed, Cochrane, Embase, CNKI, CQVIP, and Wanfang Data). Clinical research articles were screened, and animal studies as well as duplicate publications were excluded. Muscle strength and sensory recovery were considered to be effective only when the scores on the United Kingdom Medical Research Council scale were equal to or higher than M3 and S3, respectively. RESULTS: The overall ipsilateral recipient nerve recovery rates were as follows: the efficiency rate for muscle strength recovery after CC7 nerve transfer was 0.57 (95% confidence interval [CI]: 0.48-0.66) and for sensory recovery was 0.52 (95% CI: 0.46-0.58). When the recipient nerve was the median nerve, the efficiency rate for muscle strength recovery was 0.50 (95% CI: 0.39-0.61) and for sensory was 0.56 (95% CI: 0.50-0.63). When the recipient nerve was the musculocutaneous nerve and the radial nerve, the efficiency rate for muscle strength recovery was 0.74 (95% CI: 0.65-0.82) and 0.50 (95% CI: 0.31-0.70), respectively. CONCLUSIONS: Transfer of CC7 nerves to musculocutaneous nerves leads to the best results. CC7 is a reliable donor nerve, which can be safely used for upper limb function reconstruction, especially for entirely BPAI. When modifying procedures, musculocutaneous nerves and median nerve can be combined as recipient nerves.

AMP-activated protein kinase mediates the effects of lipoprotein-associated phospholipase A2 on endothelial dysfunction in atherosclerosis.

The present study aimed to investigate the effects of lipoprotein-associated phospholipase A2 (Lp-PLA2) on endothelial dysfunction in an in vitro cell model of atherosclerosis, and to determine whether AMP-activated protein kinase (AMPK) mediates the effects of Lp-PLA2 on endothelial dysfunction. A total of 392 patients with coronary artery disease (CAD), including various sub-conditions, were recruited, and the plasma concentrations of Lp-PLA2 were evaluated. In addition, an in vitro model of atherosclerosis was established by exposing human umbilical vein endothelial cells (HUVECs) to oxidized low-density lipoprotein (oxLDL). SB-435495 was used to inhibit Lp-PLA2, and compound C was used to suppress AMPK expression. Lp-PLA2, AMPKα and phosphorylated-AMPKα (T172) expression in HUVECs were evaluated using western blot analysis. The concentrations of nitric oxide (NO), endothelin 1 (ET-1), intercellular adhesion molecule 1 (ICAM-1) and platelet/endothelial cell adhesion molecule 1 (PECAM-1) in cell culture supernatant were determined using commercially available ELISA kits. MTT assays were employed to indicate changes in cell viability. The current study found the plasma Lp-PLA2 levels were elevated in the CAD patients with stable angina pectoris, unstable angina pectoris, acute coronary syndromes and acute myocardial infarction, compared with a healthy control population. In addition, the in vitro results showed that Lp-PLA2 expression levels were elevated in oxLDL-exposed HUVECs. Lp-PLA2 suppression could increase cell viability, induce the production of NO and decrease the secretion of ET-1, in addition to suppressing the expression of cell adhesion molecules, including ICAM-1 and PECAM-1 in oxLDL-exposed HUVECs. The expression of AMPKα and phosphorylated-AMPKα (T172) was regulated by Lp-PLA2, and AMPK suppression was able to reverse the effects of Lp-PLA2 with regard to cell viability, endothelial vasorelaxation capacity and the secretion of adhesion molecules in oxLDL-exposed HUVECs. In conclusion, the present study provides initial evidence that Lp-PLA2 is able to cause endothelial dysfunction in an in vitro model of atherosclerosis, and the effects of Lp-PLA2 on endothelial dysfunction was at least partially a result of the downregulation of AMPKα, thus contributing to the progression of atherosclerosis.

Phytoremediation potential of transplanted bare-root seedlings of trees for lead/zinc and copper mine tailings.

Selecting plant species that can overcome unfavorable conditions and increase the recovery of degraded mined lands remains a challenge. A pot experiment was conducted to evaluate the feasibility of using transplanted tree seedlings for the phytoremediation of lead/zinc and copper mine tailings. One-year-old bare-root of woody species (Rhus chinensis Mill, Quercus acutissima Carruth, Liquidambar formosana Hance, Vitex trifolia Linn. var. simplicifolia Cham, Lespedeza cuneata and Amorpha fruticosa Linn) were transplanted into pots with mine tailings and tested as potential metal-tolerant plants. Seedling survival, plant growth, root trait, nutrient uptake, and metal accumulation and translocation were assessed. The six species grew in both tailings and showed different tolerance level. A. fruticosa was highly tolerant of Zn, Pb and Cu, and grew normally in both tailings. Metal concentrations were higher in the roots than in the shoots of the six species. All of the species had low bioconcentration and translocation factor values. However, R. chinensis and L. formosana had significantly higher translocation factor values for Pb (0.88) and Zn (1.78) than the other species. The nitrogen-fixing species, A. fruticosa, had the highest tolerance and biomass production, implying that it has great potential in the phytoremediation of tailing areas in southern China.

Synergistic anticancer properties of docosahexaenoic acid and 5-fluorouracil through interference with energy metabolism and cell cycle arrest in human gastric cancer cell line AGS cells.

AIM: To explore the synergistic effect of docosahexaenoic acid (DHA)/5-fluorouracil (5-FU) on the human gastric cancer cell line AGS and examine the underlying mechanism. METHODS: AGS cells were cultured and treated with a series of concentrations of DHA and 5-FU alone or in combination for 24 and 48 h. To investigate the synergistic effect of DHA and 5-FU on AGS cells, the inhibition of cell proliferation was determined by MTT assay and cell morphology. Flow cytometric analysis was also used to assess cell cycle distribution, and the expression of mitochondrial electron transfer chain complexes (METCs) I, II and V in AGS cells was further determined by Western blot analysis. RESULTS: DHA and 5-FU alone or in combination could markedly suppress the proliferation of AGS cells in a significant time and dose-dependent manner. DHA markedly strengthened the antiproliferative effect of 5-FU, decreasing the IC50 by 3.56-2.15-fold in an apparent synergy. The morphological changes of the cells were characterized by shrinkage, cell membrane blebbing and decreased adherence. Cell cycle analysis showed a shift of cells into the G0/G1 phase from the S phase following treatment with DHA or 5-FU (G0/G1 phase: 30.04% ± 1.54% vs 49.05% ± 6.41% and 63.39% ± 6.83%, respectively, P < 0.05; S phase: 56.76% ± 3.14% vs 34.75% ± 2.35% and 25.63% ± 2.21%, respectively, P < 0.05). Combination treatment of DHA and 5-FU resulted in a significantly larger shift toward the G0/G1 phase and subsequent reduction in S phase (G0/G1 phase: 69.06% ± 2.63% vs 49.05% ± 6.41% and 63.39% ± 6.83%, respectively, P < 0.05; S phase: 19.80% ± 4.30% vs 34.75% ± 2.35% and 25.63% ± 2.21%, respectively, P < 0.05). This synergy was also reflected in the significant downregulation of the expression of METCs in AGS cells. CONCLUSION: Synergistic anticancer properties of DHA and 5-FU may involve interference with energy production of AGS cells via downregulation of METCs and cell cycle arrest.

Direct Coaptation of the Phrenic Nerve With the Posterior Division of the Lower Trunk to Restore Finger and Elbow Extension Function in Patients With Total Brachial Plexus Injuries.

BACKGROUND: To overcome the mismatch in nerve sizes in phrenic nerve transfer to the radial nerve for elbow and finger extension reanimation for patients with total brachial plexus injuries (TBPI), a selective neurotization procedure was designed. OBJECTIVE: To investigate the long-term results of phrenic nerve transfer to the posterior division of the lower trunk with direct coaptation in restoring elbow and finger extension after TBPI. METHODS: Phrenic nerve was transferred to and directly coapted with the posterior division of the lower trunk in 27 patients with TBPI. Seven patients were <18 years old (adolescent group), and the remaining 20 patients ≥18 years (adult group). RESULTS: Postoperative mean follow-up period was 54 ± 9 months (range, 48-85 months). The motor function attained M3 or greater in 81.5% of patients for elbow extension and in 48% of patients for finger extension. The percentage of patients who regained M3 or greater muscle power of finger extension in the adolescent group and the adult group was 71.4%, and 40%, respectively. Meanwhile, 85.7% in the adolescent group and 80% in the adult group achieved M3 or greater muscle power of elbow extension. There were no significant differences between the 2 groups. The elbow extension and finger extension were synchronous contractions and did not become independent of respiratory effort. CONCLUSION: This procedure simultaneously and effectively restores the function of elbow and finger extension in patients after TBPI. However, the patients could not do elbow and finger extension separately.

Donor-antigen Inoculation in the Testis Promotes Skin Allograft Acceptance Induced by Conventional Costimulatory Blockade via Induction of CD8 + CD122+ and CD4 + CD25+ Regulatory T Cells.

BACKGROUND: Immune responses are somewhat suppressed in immune privileged sites, including the testes, which provide a preexisting opportunity to prolong allograft survival. Previous studies have shown that intratesticular islet allografts enjoy extended survival even without any immunosuppression. However, it is unknown if testicular immune privilege can be exploited to prolong the survival of a solid allograft, including the skin, because it is impractical to implant a solid tissue in human testes. METHODS: To immunize recipient mice, splenocytes from BALB/c mice were injected into the testis of C57BL/6 recipients 1 week before skin transplantation. CD8 + CD122+ and CD4 + FoxP3+ regulatory T [Treg] cells were quantified by fluorescence-activated cell sorting. RESULTS: Although donor-antigen inoculation alone did not delay skin allograft rejection, it significantly extended the allograft survival when combined with CD40/CD40L or B7/CD28 costimulatory blockade and further induced long-term skin allograft acceptance when both costimulatory pathways were blocked. Similarly, donor-antigen inoculation suppressed alloreactive T cell proliferation in draining lymph nodes of skin recipients in the presence of the same costimulatory blockade. Interestingly, donor-antigen inoculation via intratesticular injection increased CD8 + CD122+, but not CD4 + FoxP3+, Treg numbers after transplantation. However, both CD8 + CD122+ and CD4 + CD25+ Treg cells induced by donor-antigen inoculation and the costimulatory blockade were more potent in suppression than that induced without the inoculation. Depletion of CD8+ or CD25+ T cells largely abrogated long-term skin allograft survival induced by donor-antigen inoculation and the costimulatory blockade. CONCLUSIONS: Intratesticular inoculation with donor antigens promotes long-term skin allograft survival induced by conventional costimulatory blockade via the induction of both CD8 + CD122+ and CD4 + CD25+ Treg cells.

ß-elemene enhances the radiosensitivity of gastric cancer cells by inhibiting Pak1 activation.

AIM: To explore the potential of ß-elemene as a radiosensitizer for gastric cancer cells and the underlying mechanisms. METHODS: SGC7901, MKN45, MKN28, N87, and AGS human gastric cancer cell lines were used to screen for radioresistant gastric cancer cell lines. A 3-(4,5-dimeth-ylthiazol-2-yl)-2,5-diphenyltetrazolium (MTT) assay was used to determine the effects of ß-elemene and IPA-3 on cell viability in MKN45 and SGC7901 gastric cancer cell lines. A clonogenic survival assay and annexin V-FITC/PI apoptosis detection assay were used to evaluate cellular radiosensitivity and radiation-induced cell death, respectively. A proteomic method, isobaric tags for relative and absolute quantitation (iTRAQ), was employed to screen the proteins regulated by ß-elemene pretreatment prior to ionizing radiation (IR) in SGC7901 gastric cancer cell line. IPA-3 was used as a specific small molecule inhibitor of p21-activated protein kinase 1 (Pak1) to target Pak1 signaling. Protein levels of PAK1IP1 (p21-activated protein kinase-interacting protein 1), total Pak1 (t-Pak1), phospho-Pak1 (T423), phospho-ERK1/2 (Thr202/Tyr204), and cleaved caspase-3 (17 kDa) were assessed by western blotting. RESULTS: MKN45 and SGC7901 gastric cancer cell lines were relatively more resistant to IR. ß-elemene pretreatment decreased clonogenic survival following IR in MKN45 and SGC7901 gastric cancer cell lines. Additionally, ß-elemene pretreatment prior to IR increased radiation-induced cell death compared with IR alone in MKN45 (10.4% ± 0.9% vs 34.8% ± 2.8%, P < 0.05) and SGC7901 (11.6% ± 0.9% vs 46.7% ± 5.2%, P < 0.05) human gastric cancer cell lines, respectively, consistent with the level of cleaved caspase-3 (17 kDa). Through iTRAQ analysis and western blot validation, we found that ß-elemene upregulated PAK1IP1 and downregulated phospho-Pak1 (T423) and phospho-ERK1/2 in SGC7901 gastric cancer cells. IR increased the level of phospho-Pak1 (T423). Pretreatment with ß-elemene decreased radiation-induced Pak1 and ERK1/2 phosphorylation. Inhibition of Pak1 using IPA-3 decreased clonogenic survival following IR. In addition, IPA-3 increased radiation-induced cell death in MKN45 (13.4% ± 0.3% vs 26.6% ± 1.0%, P < 0.05) and SGC7901 (16.0% ± 0.6% vs 37.3% ± 1.7%, P < 0.05) gastric cancer cell lines, respectively, consistent with the level of cleaved caspase-3 (17 kDa). Western blotting showed that IPA-3 decreased radiation-induced Pak1 and ERK1/2 phosphorylation. CONCLUSION: This is the first demonstration that ß-elemene enhances radiosensitivity of gastric cancer cells, and that the mechanism involves inhibition of Pak1 signaling.

Ischemic preconditioning ameliorates intestinal injury induced by ischemia-reperfusion in rats.

AIM: To evaluate preventative effects of ischemic preconditioning (IP) in a rat model of intestinal injury induced by ischemia-reperfusion (IR). METHODS: Male Sprague-Dawley rats (250-300 g) were fasted for 24 h with free access to water prior to the operation. Eighteen rats were randomly divided into three experimental groups: S group (n = 6), rats were subjected to isolation of the superior mesenteric artery (SMA) for 40 min, then the abdomen was closed; IR group (n = 6), rats were subjected to clamping the SMA 40 min, and the abdomen was closed followed by a 4-h reperfusion; IP group (n = 6) rats underwent three cycles of 5 min ischemia and 5 min reperfusion, then clamping of the SMA for 40 min, then the abdomen was closed and a 4-h reperfusion followed. All animals were euthanized by barbiturate overdose (150 mg/kg pentobarbital sodium, i.v.) for tissue collection, and the SMA was isolated via median abdominal incision. Intestinal histologic injury was observed. Malondialdehyde (MDA), myeloperoxidase (MPO) and tumor necrosis factor (TNF)-α concentrations in intestinal tissue were measured. Intercellular adhesion molecule (ICAM)-1 and vascular cell adhesion molecule (VCAM)-1 expression, as well as nuclear factor (NF)-κB activity and expression in intestinal tissue were also determined. RESULTS: Compared with the IR group, IP reduced IR-induced histologic injury of the intestine in rats (2.00 ± 0.71 vs 3.60 ± 0.84, P < 0.05). IP significantly inhibited the increase in MDA content (5.6 ± 0.15 µmol/L vs 6.84 ± 0.18 µmol/L, P < 0.01), MPO activity (0.13 ± 0.01 U/L vs 0.24 ± 0.01 U/L, P < 0.01), and TNF-α levels (7.79 ± 2.35 pg/mL vs 10.87 ± 2.48 pg/mL, P < 0.05) in the intestinal tissue of rats. IP also markedly ameliorated the increase in ICAM-1 (204.67 ± 53.27 vs 353.33 ± 45.19, P < 0.05) and VCAM-1 (256.67 ± 58.59 vs 377.33 ± 41.42, P < 0.05) protein expression in the intestinal tissues. Additionally, IP remarkably decreased NF-κB activity (0.48 ± 0.16 vs 0.76 ± 0.22, P < 0.05) and protein expression (320.23 ± 38.16 vs 520.76 ± 40.53, P < 0.01) in rat intestinal tissue. CONCLUSION: IP may protect against IR-induced intestinal injury by attenuation of the neutrophil-endothelial adhesion cascade via reducing ICAM-1 and VCAM-1 expression and TNF-α-induced NF-κB signaling pathway activity.

Contralateral C7 Nerve Transfer with Direct Coaptation to Restore Lower Trunk Function After Traumatic Brachial Plexus Avulsion Injuries: Surgical Technique.

INTRODUCTION: We describe a new technique for treating traumatic brachial plexus avulsion injury with a contralateral C7 nerve transfer with direct coaptation that shortens the time to muscle reinnervation. STEP 1 EXPLORE THE INJURED BRACHIAL PLEXUS: Explore the brachial plexus carefully and confirm the nerve-root avulsion injuries from C7 to T1. STEP 2 HARVEST THE CONTRALATERAL C7 NERVE: Dissect the divisions of the contralateral C7 nerve root, divide the nerve at the junction between the divisions and cords, and mobilize it proximally. STEP 3 CREATE THE PRESPINAL ROUTE: Create the prespinal route to guide the contralateral C7 nerve to the injured side. STEP 4 HUMERAL SHORTENING OSTEOTOMY: If the contralateral C7 nerve does not reach the injured lower trunk, perform a humeral shortening osteotomy, generally with <5 cm of shortening in adults. STEP 5 NEURORRHAPHY: Suture one end of the sural nerve together with the medial antebrachial cutaneous nerve to the musculocutaneous nerve; anastomose the remainder of the contralateral C7 nerve directly with the lower trunk. STEP 6 POSTOPERATIVE CARE: Use a prefabricated brace to hold the head in the neutral position and immobilize the injured limb for six weeks. RESULTS: We evaluated the results of the technique in a study of seventy men and five women with a mean age (and standard deviation) of 28 ± 10 years (range, ten to fifty-three years).IndicationsContraindicationsPitfalls & Challenges.

Contralateral C7 nerve transfer with direct coaptation to restore lower trunk function after traumatic brachial plexus avulsion.

BACKGROUND: Contralateral C7 nerve transfer to the median nerve has been used in an attempt to restore finger flexion in patients with total brachial plexus avulsion injury. However, the results have not been satisfactory mainly because of the requirement to use a long bridging nerve graft, which causes an extended nerve regeneration process and irreversible muscle atrophy. A new procedure involving contralateral C7 nerve transfer via a modified prespinal route and direct coaptation with the injured lower trunk is presented here. METHODS: Contralateral C7 nerve transfer via the modified prespinal route and direct coaptation with the injured lower trunk was performed in seventy-five patients with total brachial plexus avulsion injury. Thirty-five required humeral shortening osteotomy (3 to 4.5 cm) in order to accomplish the direct coaptation. The contralateral C7 nerve was also transferred to the musculocutaneous nerve through the bridging medial antebrachial cutaneous nerve arising from the lower trunk in forty-seven of the seventy-five patients. Recovery of finger, wrist, and elbow flexion was evaluated with use of the modified British Medical Research Council muscle grading system. RESULTS: The mean follow-up period (and standard deviation) was 57 ± 6 months (range, forty-eight to seventy-eight months). Motor function with a grade of M3+ or greater was attained in 60% of the patients for elbow flexion, 64% of the patients for finger flexion, 53% of the patients for thumb flexion, and 72% of the patients for wrist flexion. CONCLUSIONS: Contralateral C7 nerve transfer via a modified prespinal route and direct coaptation with the injured lower trunk decreases the distance for nerve regeneration in patients with total brachial plexus avulsion injury. There was satisfactory recovery of finger flexion and wrist flexion in this series. In addition, contralateral C7 nerve transfer was successfully used to repair two different target nerves: the lower trunk and the musculocutaneous nerve.