[Xanthoceras sorbifolium Bunge flower extract inhibits benign prostatic hyperplasia in rats].

OBJECTIVE: To assess the inhibitory effect of the extract of Xanthoceras sorbifolium Bunge flower against benign prostatic hyperplasia (BPH) and explore its possible mechanism. METHODS: MTT assay was used to examine the effect of the extract of Xanthoceras sorbifolium Bunge flower on proliferation of benign prostatic hyperplasia cells (BPH-1), and cell apoptosis and cell cycle changes following the treatment were analyzed using annexin V/PI double staining and flow cytometry. The protein expression levels of Bcl-2, Bax, caspase-3, PI3K and AKT in the treated cells were detected using Western blotting. A rat model of BPH established by subcutaneous injection of testosterone propionate was treated with the flower extract for 28 days, and pathological changes in the prostate tissue were observed with HE staining. The protein expression levels of Bcl-2, Bax, caspase3 and PI3K/AKT in the prostate tissue were detected with Western blotting. RESULTS: Within the concentration range of 125-1000 µg/mL, the flower extract of Xanthoceras sorbifolium Bunge significantly inhibited the proliferation of BPH-1 cells and caused obvious cell cycle arrest at G0/G1 phase; the apoptotic rate of the cells was positively correlated with the concentration of the flower extract (P < 0.05). Bcl-2, p-PI3K and p-AKT expression levels were significantly down-regulated and Bax and caspase-3 expression levels were significantly increased in the cells after treatment with the flowers extract (P < 0.05). In the rat models of BPH, the rats treated with the flowers extract at moderate and high doses showed obviously decreased expressions of p-AKT and Bcl-2 and an increased expression of Bax in the prostate tissue; a significantly lowered p-AKT expression was observed in the prostate tissue of rats receiving the low-dose treatment (P < 0.05). CONCLUSION: The flower extract of Xanthoceras sorbifolium Bunge has a inhibitory effect on BPH both in vitro and in rats, suggesting its potential value in the development of medicinal plant preparations for treatment of BPH.

Protective effects of curcumin against mercury-induced hepatic injuries in rats, involvement of oxidative stress antagonism, and Nrf2-ARE pathway activation.

Mercury (Hg) represents a ubiquitous environmental heavy metal that could lead to severe toxic effects in a variety of organs usually at a low level. The present study focused on the liver oxidative stress, one of the most important roles playing in Hg hepatotoxicity, by evaluation of different concentrations of mercuric chloride (HgCl2) administration. Moreover, the protective potential of curcumin against Hg hepatotoxic effects was also investigated. Eighty-four rats were randomly divided into six groups for a three-days experiment: control, dimethyl sulfoxide control, HgCl2 treatment (0.6, 1.2, and 2.4 mg kg-1 day-1), and curcumin pretreatment (100 mg kg-1 day-1) groups. Exposure of HgCl2 resulted in acute dose-dependent hepatotoxic effects. Administration of 2.4 mg kg-1 HgCl2 significantly elevated total Hg, nonprotein sulfhydryl, reactive oxygen species formation, malondialdehyde, apoptosis levels, serum lactate dehydrogenase, and alanine transaminase activities, with an impairment of superoxide dismutase and glutathione peroxidase in the liver. Moreover, HgCl2 treatment activated nuclear factor-E2-related factor 2-antioxidant response element (Nrf2-ARE) signaling pathway in further investigation, with a significant upregulation of Nrf2, heme oxygenase-1, and γ-glutamylcysteine synthetase heavy subunit expression, relative to control. Pretreatment with curcumin obviously prevented HgCl2-induced liver oxidative stress, which may be due to its free radical scavenging or Nrf2-ARE pathway-inducing properties. Taking together these data suggest that curcumin counteracts HgCl2 hepatotoxicity through antagonizing liver oxidative stress.

Risk factors for pulmonary hypertension in patients receiving maintenance peritoneal dialysis

We investigated the risk factors for pulmonary hypertension (PH) in patients receiving maintenance peritoneal dialysis (MPD). A group of 180 end-stage renal disease patients (124 men and 56 women; mean age: 56.43±8.36) were enrolled in our study, which was conducted between January 2009 and June 2014. All of the patients received MPD treatment in the Dialysis Center of the Second Affiliated Hospital of Soochow University. Clinical data, laboratory indices, and echocardiographic data from these patients were collected, and follow-ups were scheduled bi-monthly. The incidence and relevant risk factors of PH were analyzed. The differences in measurement data were compared by t-test and enumeration data were compared with the χ2 test. Among the 180 patients receiving MPD, 60 were diagnosed with PH. The remaining 120 were regarded as the non-PH group. Significant differences were observed in the clinical data, laboratory indices, and echocardiographic data between the PH and non-PH patients (all P<0.05). Furthermore, hypertensive nephropathy patients on MPD showed a significantly higher incidence of PH compared with non-hypertensive nephropathy patients (P<0.05). Logistic regression analysis showed that the proportion of internal arteriovenous fistula, C-reactive protein levels, and ejection fraction were the highest risk factors for PH in patients receiving MPD. Our study shows that there is a high incidence of PH in patients receiving MPD and hypertensive nephropathy patients have an increased susceptibility to PH.

Risk factors for pulmonary hypertension in patients receiving maintenance peritoneal dialysis.

We investigated the risk factors for pulmonary hypertension (PH) in patients receiving maintenance peritoneal dialysis (MPD). A group of 180 end-stage renal disease patients (124 men and 56 women; mean age: 56.43±8.36) were enrolled in our study, which was conducted between January 2009 and June 2014. All of the patients received MPD treatment in the Dialysis Center of the Second Affiliated Hospital of Soochow University. Clinical data, laboratory indices, and echocardiographic data from these patients were collected, and follow-ups were scheduled bi-monthly. The incidence and relevant risk factors of PH were analyzed. The differences in measurement data were compared by t-test and enumeration data were compared with the χ2 test. Among the 180 patients receiving MPD, 60 were diagnosed with PH. The remaining 120 were regarded as the non-PH group. Significant differences were observed in the clinical data, laboratory indices, and echocardiographic data between the PH and non-PH patients (all P<0.05). Furthermore, hypertensive nephropathy patients on MPD showed a significantly higher incidence of PH compared with non-hypertensive nephropathy patients (P<0.05). Logistic regression analysis showed that the proportion of internal arteriovenous fistula, C-reactive protein levels, and ejection fraction were the highest risk factors for PH in patients receiving MPD. Our study shows that there is a high incidence of PH in patients receiving MPD and hypertensive nephropathy patients have an increased susceptibility to PH.

Black cohosh improves objective sleep in postmenopausal women with sleep disturbance.

OBJECTIVE: Sleep problems are prominent after menopause. The aim of our study was to look into the effect of black cohosh on both objective and subjective sleep in early postmenopausal women with sleep complaints. METHODS: We performed a randomized, double-blind and placebo-controlled research during a 6-month period. Forty-eight postmenopausal women aged 45-60 years with sleep disturbance were enrolled and received daily administration of either black cohosh or placebo. Polysomnography and the Pittsburg Sleep Quality Index (PSQI) were performed at the initiation and termination of the study, as well as the Menopause-specific Quality of Life questionnaire and estradiol and follicle stimulating hormone tests. Liver and renal functions and breast and pelvic ultrasound were set as safety measures, carried out every 3 months. RESULTS: Seventy-six women were interviewed, of whom 42 women completed the whole trial. Compared with placebo, black cohosh treatment led to significant polysomnographic changes, including increased sleep efficiency and decreased wake after sleep onset (WASO) duration, and tended to improve PSQI with a medium effect size. On average, 15.8% of WASO duration was reduced in the black cohosh group. Vasomotor and physical domains of life quality were improved compared with placebo. Safety measures did not yield any adverse event assigned to black cohosh. CONCLUSIONS: In early postmenopausal women with a major sleep complaint, black cohosh effectively improved sleep and might be a safe measure in managing menopausal sleep disturbance.

The protective role of tea polyphenols against methylmercury-induced neurotoxic effects in rat cerebral cortex via inhibition of oxidative stress.

Methylmercury (MeHg) is a ubiquitous environmental contaminant that could induce oxidative stress and an indirect glutamate (Glu)-mediated excitotoxicity. However, the underlying mechanisms through which MeHg affects the central nervous system have not been fully elucidated, and little has been known of the interaction between oxidative stress and Glu dyshomeostasis in MeHg neurotoxicity. Therefore, rats were administrated with different MeHg concentrations (0, 4, and 12 µmol/kg) to evaluate the neurotoxic effects in cerebral cortex. Moreover, we have investigated the neuroprotective role of tea polyphenols (TP), a natural antioxidant that has a formidable free radical scavenge ability, against MeHg-induced neurotoxicity. Eighty rats were randomly divided into five groups: control, TP control, MeHg-treated (4 and 12 µmol/kg), and TP pretreated (1 mmol/kg). Administration of MeHg at 12 µmol/kg for 4 weeks significantly increased total Hg and ROS levels in cerebral cortex. In addition, MeHg reduced non-enzymatic (non-protein sulfhydryl) and enzymatic (SOD and GSH-Px) antioxidants, up-regulated Nrf2, HO-1, and γ-GCS expression. Moreover, MeHg-induced ROS over-production appeared to inhibit the activities of GS, down-regulated GLAST and GLT-1 expression in cerebral cortex. Pretreatment with TP at a dose of 1 mmol/kg significantly prevented MeHg-induced oxidative stress and Glu uptake/metabolism disorders in cerebral cortex. In conclusion, the results suggested that oxidative stress resulting from excessive ROS formation plays a critical role in MeHg neurotoxicity. TP possesses the ability to attenuate MeHg-induced neurotoxic effects through its antioxidative properties.

Ginsenoside Rh2 promotes nonamyloidgenic cleavage of amyloid precursor protein via a cholesterol-dependent pathway.

Ginsenoside Rh2 (Rh2) is a ginseng derivative used in Chinese traditional medicine. We investigated whether Rh2 can help prevent Alzheimer's disease symptoms and examined underlying mechanisms. We injected Rh2 into tg2576 Alzheimer's disease model mice and looked for behavioral improvement and senile plaque reduction in brain slices. We measured amyloid precursor protein (APP) metabolism species changes, amyloid beta40 and 42 levels and ß, γ secretase activity in primary hippocampal neurons. By living cell staining, we detected surface and endocytosed APP. We also measured cholesterol and lipid rafts in primary neurons. Rh2 treatment significantly improved learning and memory performance at 14 months of age; it also reduced brain senile plaques at this age. Based on in vitro experiments, we found that Rh2 treatment increased soluble APPα (sAPPα) levels, increased CTFα/ß ratios, and reduced amyloid beta 40 and 42 concentrations. Surface APP levels dramatically increased. Based on living cell staining, we found that Rh2 inhibited APP endocytosis. Based on lipid removal and reload experiments, we found that Rh2 can modulate APP by reducing cholesterol and lipid raft levels. We concluded that Rh2 improves learning and memory function in Alzheimer's disease model mice, and that this improvement is accomplished by reducing amyloid beta secretion and APP endocytosis, which in turn is achieved by reducing cholesterol and lipid raft concentrations.

Proteomic Analysis of RBP4/Vitamin A in Children with Cleft Lip and/or Palate.

Cleft of the lip and/or palate (CLP) is one of the most common congenital craniofacial defects. Non-syndromic CLP (NSCLP) is a multifactorial disease influenced by the interaction of genetic and environmental factors. However, there are few studies reporting on the developmental or metabolic status of babies with NSCLP after birth. In our study, we sought to identify and evaluate the differential expression of serum protein profiles in NSCLP children and unaffected babies. Thus, a 'shotgun proteomics' approach was first used to analyze the plasma proteome of 13 children with NSCLP and 10 control children, aged 2 to 3.5 years. In total, more than 300 proteins were identified in the serum sample. With gene ontology (GO) analysis, we detected many differentially expressed proteins that could be related to NSCLP, including those involved in lipoprotein metabolism, insulin-like growth-factor-related processes, and so on, especially the proteins involved in retinol transport. Retinol binding protein 4 (RBP4), one protein of the retinol transport category, was significantly decreased in the NSCLP group. Thus, serum vitamin A levels were further determined by high-performance liquid chromatography (HPLC). A significant difference (p < .01) was also found in vitamin A concentrations, consistent with the trend of RBP4. Our results indicated that reduced levels of RBP4 and vitamin A were related to newborns with NSCLP and should thus receive more attention. These results also suggest that vitamin A supplementation might be necessary at an early stage.

Glycidyl fatty acid esters in food by LC-MS/MS: method development.

An improved method based on liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the analysis of glycidyl fatty acid esters in oils was developed. The method incorporates stable isotope dilution analysis (SIDA) for quantifying the five target analytes: glycidyl esters of palmitic (C16:0), stearic (C18:0), oleic (C18:1), linoleic (C18:2) and linolenic acid (C18:3). For the analysis, 10 mg sample of edible oil or fat is dissolved in acetone, spiked with deuterium labelled analogs of glycidyl esters and purified by a two-step chromatography on C18 and normal silica solid phase extraction (SPE) cartridges using methanol and 5% ethyl acetate in hexane, respectively. If the concentration of analytes is expected to be below 0.5 mg/kg, 0.5 g sample of oil is pre-concentrated first using a silica column. The dried final extract is re-dissolved in 250 µL of a mixture of methanol/isopropanol (1:1, v/v), 15 µL is injected on the analytical C18 LC column and analytes are eluted with 100% methanol. Detection of target glycidyl fatty acid esters is accomplished by LC-MS/MS using positive ion atmospheric pressure chemical ionization operating in Multiple Reaction Monitoring mode monitoring 2 ion transitions for each analyte. The method was tested on replicates of a virgin olive oil which was free of glycidyl esters. The method detection limit was calculated to be in the range of 70-150 µg/kg for each analyte using 10 mg sample and 1-3 µg/kg using 0.5 g sample of oil. Average recoveries of 5 glycidyl esters spiked at 10, 1 and 0.1 mg/kg were in the range 84% to 108%. The major advantage of our method is use of SIDA for all analytes using commercially available internal standards and detection limits that are lower by a factor of 5-10 from published methods when 0.5 g sample of oil is used. Additionally, MS/MS mass chromatograms offer greater specificity than liquid chromatography-mass spectrometry operated in selected ion monitoring mode. The method will be applied to the survey of glycidyl fatty acid esters in food products on the Canadian market.

Release of paclitaxel from polylactide-co-glycolide (PLGA) microparticles and discs under irradiation.

Paclitaxel is a promising anti-cancer drug as well as a radiosensitizer for chemotherapy and radiotherapy applications. Because of the poor solubility of paclitaxel in water and most pharmaceutical reagents, it is usually formulated with an adjuvant called Cremophor EL, which causes severe side effects. This work develops new dosage forms of paclitaxel for controlled release application, which do not require the adjuvant and, thus, can avoid its associated side effects. Paclitaxel was encapsulated into the PLGA matrix with various additives such as polyethylene glycol (PEG), isopropyl myristate (IPM) and d-alpha tocopheryl polyethylene glycol (Vitamin E TPGS). These additives were used to enhance the release rate of paclitaxel from the polymer matrix. Spray-drying and an hydraulic press were used to prepare paclitaxel-PLGA microspheres and discs. The microspheres and discs were given different irradiation doses to investigate their effects on the surface morphology (characterized by SEM, AFM and XPS) and in vitro release properties. There seems to be a small effect of the ionizing radiation on various formulations. Although the irradiation did not cause observable changes on the morphology of the polymer matrix, the release rate can be enhanced by a few per cent. It was found that PEG has the highest enhancement effect for release rate among all the additives investigated in this study.

A novel controlled release formulation for the anticancer drug paclitaxel (Taxol): PLGA nanoparticles containing vitamin E TPGS.

Paclitaxel (Taxol) is one of the best antineoplastic drugs found from nature in the past decades. Like many other anticancer drugs, there are difficulties in its clinical administration due to its poor solubility. Therefore an adjuvant called Cremophor EL has to be employed, but this has been found to cause serious side-effects. However, nanoparticles of biodegradable polymers can provide an ideal solution to the adjuvant problem and realise a controlled and targeted delivery of the drug with better efficacy and fewer side-effects. The present research proposes a novel formulation for fabrication of nanoparticles of biodegradable polymers containing d-alpha-tocopheryl polyethylene glycol 1000 succinate (vitamin E TPGS or TPGS) to replace the current method of clinical administration and, with further modification, to provide an innovative solution for oral chemotherapy. In the modified solvent extraction/evaporation technique employed in this research, the emulsifier/stabiliser/additive and the matrix material can play a key role in determining the morphological, physicochemical and pharmaceutical properties of the produced nanoparticles. We found that vitamin E TPGS could be a novel surfactant as well as a matrix material when blended with other biodegradable polymers. The nanoparticles composed of various formulations and manufactured under various conditions were characterised by laser light scattering (LLS) for size and size distribution, scanning electron microscopy (SEM) and atomic force microscopy (AFM) for morphological properties, X-ray photoelectron spectroscopy (XPS) for surface chemistry and differential scanning calorimetry (DSC) for thermogram properties. The drug encapsulation efficiency (EE) and the drug release kinetics under in vitro conditions were measured by high performance liquid chromatography (HPLC). It was concluded that vitamin E TPGS has great advantages for the manufacture of polymeric nanoparticles for controlled release of paclitaxel and other anti-cancer drugs. Nanoparticles of nanometer size with narrow distribution can be obtained. A drug encapsulation efficiency as high as 100% can be achieved and the release kinetics can be controlled.

Relationship of blood trace elements to liver damage, nutritional status, and oxidative stress in chronic nonalcoholic liver disease.

Trace elements are involved in chronic liver diseases because these elements may have a direct hepatic toxicity or may be decreased as a consequence of the impaired liver function, particularly in patients with alcoholic cirrhosis and/or malnutrition. In this study, we determined plasma and erythrocytes trace elements in 50 inpatients with nonalcoholic chronic liver disease (11 with biopsy-proven chronic hepatitis, 39 with cirrhosis [16 in stage A according to Child-Pugh criteria, 23 Child B+C]), and in a control group of 10 healthy subjects by the proton induced x-ray emission method. The relationship between trace element concentration and the extent of liver damage, the nutritional status (by anthropometric evaluations), and various blood markers of oxidative stress--reduced glutathione, total lipoperoxides and malonyldialdehyde--was investigated. We found that cirrhotics had a significant decrease of Fe, Zn, Se, and GSH levels in the plasma and of GSH and Se in the erythrocytes with respect to the control and chronic hepatitis groups. GSH levels were related to the degree of liver damage; a significant direct correlation was observed among Se, Zn, and GSH plasma values and between GSH and Se in the erythrocytes. The trace element decrease was, on the contrary, independent of the degree of liver function impairment and only partially affected by the nutritional status. Data indicate that liver cirrhosis, even if not alcohol related, induces a decrease of Se and Zn and that, in these patients, an oxidative stress is present, as documented by the significant correlation between Se and GSH. The plasma Br level was higher in cirrhotics with respect to the control and chronic hepatitis groups.

RAPD-PCR typing of Acinetobacter isolates from activated sludge systems designed to remove phosphorus microbiologically.

AIMS: This study investigated whether there were differences in RAPD fingerprints between already described genomic species of Acinetobacter and those from activated sludge systems. Whether plant-specific populations of acinetobacters exist was also examined. METHODS AND RESULTS: Fifty-two isolates of Acinetobacter from four biological phosphorus removal (EBPR) systems of different configurations, and the known genomic species, were characterized using RAPD-PCR, and fragments separated on agarose gels. Patterns were analysed using Gel Pro software and data analysed numerically. RAPD-PCR produced patterns suggesting that many environmental isolates differ from known genomic species. In two cases, strains from individual plants clustered closely enough together to imply that there may be plant-specific populations of acinetobacters. CONCLUSION: The data suggest that current understanding of the taxonomic status of Acinetobacter may need modifying to accommodate non-clinical isolates, as many of the clusters emerging after numerical analysis of RAPD-PCR fragments from activated sludge isolates were quite separate from the clusters containing the already described genomic species. Some evidence was also obtained from the clusters generated to support a view that particular populations of Acinetobacter may occur in individual activated sludge plants. SIGNIFICANCE AND IMPACT OF THE STUDY: These data suggest that the current understanding of the systematics of Acinetobacter, based as it is almost exclusively on clinical isolates, may need drastic revision to accommodate environmental strains. They also suggest that a re-examination of the importance and role of Acinetobacter in the activated sludge process may be appropriate.

Effect of ligustrum fruit extract on reproduction in experimental diabetic rats.

AIM: To study the effect of ligustrum fruit on spermatogenesis and blood gonadal hormones in diabetic rats. METHODS: Experimental diabetes was induced in male Wistar rats with streptozotocin. Ligustrum fruit extract was given by gastric gavage at a dose of crude drug 30 g x kg(-1) x d(-1) for 110 days. The serum gonadadotropic hormones and testosterone were determined on d 60 and testicular histology examined on d 110. RESULTS: In the control diabetic rats, the seminiferous tubules were dilated and the spermatogenic cells irregularly arranged. Spermatogenesis was arrested with the number of spermatids highly reduced and spermatozoa not observed. In the treated rats, all types of spermatogenic cells were practically normal. The serum luteinizing hormone (LH), follicle-stimulating hormone (FSH) and testosterone levels were higher in the treated than in the control rats, but the difference was insignificant. CONCLUSION: In experimental diabetic rats, ligustrum fruit extract protects the damaging effect of experimental diabetes on spermatogenesis.

[Chemical constituents from Salix oritrepha Schneid].

OBJECTIVE: To study chemical constituents of the above ground part of Salix oritrepha. METHOD: Using chromatography to isolate compounds and their structure were identified by physical, chemical and spectral techniques. RESULT: Four compounds were isolated and elucidated as beta-sitosterol, beta-sitosteryl-3-O-glucoside, friedelin, luteolin-7-O-glucoside. CONCLUSION: All of them were isolated from S. oritrepha for the first time. Friedelin from the plants of genus Salix for the first time.

Decreased bone mineral density in female patients with systemic lupus erythematosus after long-term administration of Tripterygium Wilfordii Hook. F.

OBJECTIVE: To investigate the possible bone changes in female patients with systemic lupus erythematosus (SLE) induced by long-term administration of Tripterygium Wilfordii Hook. F (TW). METHODS: 70 female SLE patients were divided into 4 groups according to their drug history: SLE disease control group, corticosteroids treatment group, TW treatment group, and both corticosteroids and TW treatment group. Bone mineral density (BMD) of the lumbar spine 2-4 and biochemical markers of bone turnover were studied. RESULTS: Long-term administration of TW could significantly decrease BMD levels in female SLE patients (P < 0.05). The patients receiving TW for more than 5 years had significantly lower BMD levels compared with those for less than 5 years. The degree of decreased BMD induced by TW was less severe compared with that of prednisone. No significant differences were observed in the biochemical markers of bone turnover among four groups (P > 0.05). CONCLUSION: Long-term administration of TW could decrease BMD levels in women. Osteoporosis may be an important problem for SLE patients treated with TW.

Establishment of a pure culture of distinct cell types from bovine placental cotyledon.

Methods are described to establish distinct cell cultures from bovine placental cotyledon. The villous tissue of the bovine placental cotyledon is collected and dissociated with 0.125% trypsin. The cells are then cultured in three different media: a serum-free medium, a growth factor supplemented medium, and a medium with 10% serum. A polygonal cell line grew out of the serum-containing medium, a fan-shaped cell line grew out of the serum-free medium, and an epitheloid cell line grew out of the growth factor supplemented medium. These cells maintained their morphology when grown in serum containing medium. The preference of distinct cells for different media in vitro reflects the in vivo physiological regulation of these cells. These distinct cultures re ideal to study the extrinsic and interactive factors in bovine placenta.

Long-term zinc and iron supplementation in children of short stature: effect of growth and on trace element content in tissues.

We evaluated the effect of one year of supplementation with iron plus zinc (12 mg/day of Fe+++ and 12.5 mg/day of Zn++), zinc alone (12.5 mg/day of Zn++) and placebo on growth and on the iron, zinc, copper and selenium tissue contents in 30 well-selected children of short stature (16 M and 14 F; 4-11 years old). Before and after supplementation, we measured the concentrations of iron, transferrin, ferritin, zinc and copper in serum, of zinc in erythrocytes and leukocytes, and of zinc, copper and selenium in hair, as well as glutathione peroxidase activity in erythrocytes. Before supplementation, ferritin and serum, erythrocyte and hair zinc contents were significantly lower than in age-matched controls, while the other measured indices were in the normal range. Iron plus zinc supplementation caused an improvement in growth rate in all subjects, i.e., the median Z-score increased from -2.22 +/- 0.45 to -0.64 +/- 0.55; (p < 0.01). In the zinc-supplemented group, only the subjects whose ferritin levels were higher than 20 ng/L before supplementation showed a similar improvement of growth rate. Iron plus zinc supplementation could be a reasonable treatment in short, prepubertal children affected by marginal zinc and iron deficiency.